bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential growth response of normal human diploid fibroblasts and in vitro transformed human fibroblasts in serum‐free defined culture medium
Authors: Namba Masayoshi , Kusaka Takashi , Fukushima Fujiko , Kimoto Tetsuo
Published: February 2007

Two neoplastic human cell lines, WI‐38 CT‐1 and SUSM‐1, which were transformed in vitro with gamma rays and 4‐nitroquinoline 1‐oxide, respectively, grew continuously in a serum‐free defined medium. The defined medium used was a 1:1 mixture by volume of Dulbecco's modified Eagle's medium and Ham's F12 (DF) supplemented with 0.1% bovine serum albumin fraction V, 10 μg/ml of transferrin, 1 μg/ml of insulin, and 5 of μg/ml oleic acid. In the case of SUSM‐1, 100 μg/ml of fetuin were added to cultures when the cells were subcultured. Under these conditions the growth rates of the two transformed human cell lines were almost the same as those in a DF medium containing 10% fetal bovine serum (FBS). In addition, the defined medium permitted the cells to grow indefinitely without a lag period when they were transferred from serum‐containing medium into this defined medium, indicating that no selection or adaptation of the cells had occurred. Interestingly, these cell lines did not require for their growth any polypeptide growth factors such as epidermal, platelet‐derived or fibroblast growth factors. On the other hand, the control WI‐38 cells stopped growing in the defined medium after about 2 divisions. Another control normal cell strain of fibroblasts derived from a human embryo showed a decreased growth rate in the defined medium as compared with that in the DF medium with 10% FBS. These results suggest that the defined medium described here is useful for the selective growth of human cells transformed in vitro after treatment with carcinogens from an untransformed cell population. In addition, the defined medium for transformed human cells should contribute to studies on their growth mechanisms.
Suggestive evidence that the highly metastatic variant ESB of the T‐cell lymphoma eb is derived from spontaneous fusion with a host macrophage
Authors: Larizza L. , Schirrmacher V. , Graf L. , Pflüger E. , Peres‐Martinez M. , Stöhr M.
Published: February 2007

Two lines of evidence are reported which suggest that the highly metastatic variant ESb of the T‐cell lymphoma Eb is derived from spontaneous fusion with a host macrophage. Firstly, ESb cells are shown to express the macrophage differentiation antigen Mac‐1 which was not found on Eb cells or on any other tumor cells tested except the macrophage tumor line Pu5. Secondly, the progression from low to high metastatic capacity could be reproduced in vitrofollowing hybridization of thioguanine‐resistant Eb cells (EbTGR) with syngeneic bone‐marrow‐derived macrophages. Two HAT medium‐selected hybrid tumor lines (Eb‐F1 and Eb‐F2) could be established. They were found to express cell surface markers of both parental lines: T lymphoid differentiation antigens from T‐lymphoma and macrophage antigens (Mac‐1, class II MHC antigens) from the normal cell fusion partner. The antigens were identified on the hybrids and subclones thereof by means of monoclonal antibodies and 3 different detection assays: cytofluorography, complement‐dependent cytotoxocity and immunoprecipitation followed by gel electrophoresis. Animals inoculated s.c. with the parental line EbTGR developed local tumors but not metastases and survived for more than 40 days. In contrast, animals inoculated similarly with Eb‐F1 or Eb‐F2 cells quickly developed metastases in visceral organs and died as early as 10‐14 days following inoculation. In many but not all respects, the in vitro‐derived Tlymphoma‐macrophage hybrids resembled the spontaneous in vivo‐derived variant ESb. These findings, together with the presence of Mac‐1 antigen on ESb cells, suggest (1) that ESb variant cells may be derived from spontaneous fusion with a host cell, most likely a macrophage and (2) that somatic cell fusion may be an important mechanism of genetic rearrangements leading to metastatic variants. The new highly metastatic tumor lines which were developed under well‐defined in vitro conditions, and their subclones, may become very useful tools for studying the contribution of specific genetic traits and of membranerelated structures to various steps of the metastatic process.
Use of the micronucleus test to monitor the effect of vitamin A, beta‐carotene and canthaxanthin on the buccal mucosa of betel nut/tobacco chewers
Authors: Stich Hans F. , Stich Waltraud , Rosin Miriam P. , Vallejera Mythyl O.
Published: February 2007

The frequency of exfoliated cells with micronuclei in buccal swabs was used to estimate the protective effect of vitamin A, beta‐carotene and canthaxanthin (4,4′‐diketo‐beta‐carotene) on the buccal mucosa of betel (areca) nut/tobacco chewers. Micronuclei were scored on exfoliated cells taken by swabbing and stained with the Feulgen reaction and fast green. The betel (areca) nut/tobacco chewers served as their own controls. Prior to the administration of vitamin A and beta‐carotene, the examined betel quid chewers had elevated frequencies of micronucleated buccal mucosa cells, averaging 4.03% ± 1.24 sd (n = 26) and 3.43% ± 1.22 sd (n = 25), respectively. The frequency of micronucleated buccal mucosa cells in non‐chewers and non‐smokers was 0.51% (n = 52). Following a 9‐week ingestion of vitamin A (150,000 IU/week) and beta‐carotene (180 mg/week in 6 capsules), the frequency of micronucleated cells decreased significantly (p<0.001) to 1.70% and 1.16%, respectively. No significant shift in the frequencies of micronucleated cells was observed following the intake of canthaxanthin (180 mg/week in 6 capsules) for 9 weeks or that of a placebo. The lack of protective activity of canthaxanthin, which is a good trapper of oxygen singlets but cannot be converted into vitamin A, suggests that vitamin A and beta‐carotene exert their inhibitory effect on the formation of micronuclei by a mechanism not involving the scavenging of free radicals. The efficacy of beta‐carotene as an inhibitor of micronucleated cell formation, the lack of toxicity, and its availability from a multitude of dietary sources should focus attention on this carotenoid as a promising chemopreventive agent.
The proliferative response of low‐density human cell cultures to tumor promoters and its relevance to carcinogenic mechanisms in vitro
Authors: Kopelovich Levy , Chou Ting‐Chao
Published: February 2007

The effects of the tumor promoter 12‐O‐tetradecanoyl phorbol‐13‐acetate, its analogues, and other tumor promoters, on the proliferation of human skin fibroblasts (SF) have been investigated. We have previously shown (Kopelovich and Bias, 1979), that TPA caused a biphasic (upward concave) dose effect in the cloning efficiency assay of normal and mutant human fibroblastic cell strains. Here we report that the biphasic dose response pattern, consisting of an inhibitory phase and a stimulatory phase, was shared by the active analogues of TPA, e.g., phorbol‐12, 13‐dibutyrate and phorbol‐12, 13‐dibenzoate. This biphasic dose effect relationship, however, was not seen with phorbol‐12, 13‐diacetate or the inactive analogues of TPA such as phorbol and 4‐O‐methyl‐12‐O‐tetradecanoyl phorbol‐13‐acetate, nor was it seen with mezerein, teleocidin, or bile‐acid derivatives of humans. An analysis of the cloning efficiency data by the median‐effect equation (Chou and Talalay, 1981) showed that in low‐density cultures both the inhibitory phase and the stimulatory phase of the dose‐effect relationships of TPA, its analogues, mezerein, and teleocidin exhibited a linear median‐effect plot and thus closely followed the basic mass‐action principle. The median‐effect plot of these data allowed quantitative determination of growth curve characteristic such as regression coefficient, slope (a measure of sigmoidicity), median‐effect concentrations such as I50 for the inhibitory effect, and A50 for the stimulatory effect (i.e., the relative potency of the analogues) and the transition point of the biphasic phenomenon in the case of the phorbol esters. In additon, we have demonstrated a relationship between the dose response effect of TPA on the proliferation of various human cells and tumor progression in vitro.
Induction of IL‐2 production, IL‐2 receptor expression and proliferation of T3− T‐PLL cells by phorbol ester
Authors: Vyth‐Dreese Florry A. , De Vries Jan E.
Published: February 2007

Leukemic T cells from the peripheral blood of a patient with T‐prolymphocytic leukemia (T‐PLL) were investigated for their potential to differentiate in vitro upon exposure to 12‐O‐tetradecanoyl‐phorbol‐13‐acetate (TPA). The T‐PLL cells, identified as typical PLL cells by nuclear morphology, were typed as E+ sIg−OKTI+3−4+6−8−11+ cells lacking reactivity with OKI‐I or OKM‐l. In addition, between 3% and 10% of the cells reacted with monoclonal antibodies against T10. In contrast to normal T cells, the T‐PLL cells could not be induced to proliferate by mitogenic lectins or alloantigens in the presence or absence of human interleukin‐l (IL‐I), interleukin‐2 (IL‐2) or allogeneic monocytes and did not produce IL‐2. They also failed to proliferate in response to TPA or TPA in the presence of phytohemagglutinin (PHA), but under these conditions T‐PLL cells secreted high levels of IL‐2 activity. Incubation in the presence of PHA + TPA or TPA for 48 h induced T‐PLL cells to become blasts exhibiting enhanced protein synthesis, and induced a 10‐fold increase in the percentage of cells reactive with monoclonal antibodies against T10. At the same time, about 15% of the cells developed receptors for IL‐2 as monitored by their reactivity with anti‐Tac monoclonal antibody. Washing of these T‐PLL cells to remove TPA resulted in the induction of proliferation upon subsequent culture in the presence of IL‐2 or in medium only. Since proliferating T‐PLL cells still failed to express T3 antigens, it was concluded that these leukemic cells represent a T‐cell differentiation stage or a T‐cell subset which can be activated via a T3‐independent pathway.
In vitro analysis of the epstein‐barr virus: Host balance in long‐term renal allograft recipients
Authors: Yao Q. Y. , Rickinson A. B. , Gaston J. S. H. , Epstein M. A.
Published: February 2007

Four indlces of the EB virus carrier state, for which quantitative in vitro assays now exist, have been monitored in 55 renal allograft recipients under long‐term immunosuppression, each patient being tested on a single occasion. By comparison with parallel data from healthy control donors, the results indicate the extent to which virus replication in the throat and virusinfected B cells in the blood are increased as a result of immunosuppression; the concrodance between these two independent indices of the level of EB virus infection in vivo, first noted with healthy donors, was again observed within this large group of patients. Immunosuppression also leads to an impairment of virus‐specific memory T‐cell responsiveness and to an increase in anti‐viral antibody titres, but the results show that the level of virus infection prevailing in any one individual cannot be inferred directly from these immunological indices of the virus: host balance. In allograft patients on stable levels of immunosuppression, virus and host appear to establish a new equilibrium. Limited prospective studies suggest that the position of this new equilibrium depends critically upon the virus: host balance prevalling in the same individuals before immunosuppression began. This may be an important consideration in identifying patients for whom immunosppression may carry a particularly high risk of developing EB virus genome‐positive lymphoma.
Herpes simplex virus type 2 and human papillomavirus type 16 in cervicitis, dysplasia and invasive cervical carcinoma
Authors: Prakash S. S. , Reeves William C. , Sisson Gary R. , Brenes Maria , Godoy Julio , Bacchetti Silvia , de Britton Rosa C. , Rawls William E.
Published: February 2007

Sera and biopsies of cervical lesions from 55 women with benign or malignant disease were analyzed for evidence of infection with herpes simplex virus type 2 (HSV‐2) or human papillomavirus (HPV). In addition, information regarding known risk factors for cervical cancer was obtained by interview. The sera were tested for HSV‐2 antibodies and the biopsies were tested for HPV or HSV DNA sequences by Southern blot hybridization. HSV‐2 sequences were detected in 2/13 (15%) invasive neoplasms and in 1/12 (7%) benign lesions. Under non‐stringent conditions of hybridization, reactions with HPV DNA were detected in biopsies of 2/17 (12%) inflammatory lesions, 6/12 (50%) intraepithelial neoplasms and 13/20 (65%) invasive neoplasms. All but one of the positive biopsies of invasive cancer, but only 4/11 biopsies of non‐invasive lesions, contained HPV‐16 DNA as determined by stringent hybridization conditions. Women with cervical cancer possessed the risk factors associated with the disease. Cigarette smoking and the presence of HPV‐16 DNA were the most prominent risk factors. No evidence of an interaction between HSV‐2 and HPV‐16 was found among the cases of invasive cervical cancer.
Endocrine sensitivity of the receptor‐positive T61 human breast carcinoma serially grown in nude mice
Authors: Brünner N. , Spang‐Thomsen M. , Poulsen H. Skovgaard , Engelholm S. A. , Nielsen A. , Vindeløv L.
Published: February 2007

A study was made of the effect of ovariectomy, 17β‐oestradiol, and tamoxifen on the oestrogen and progesterone receptor‐positive T61 human breast carcinoma grown in nude mice. The effect of the treatment was evaluated by the specific growth delay calculated on the basis of Gompertz growth curves, and by the changes in the cell cycle distribution monitored by flow cytometric DNA analysis. The results demonstrated that both oestradiol and tamoxifen induced a temporary growth delay, whereas ovariectomy of the host had no effect on the growth of the tumour. The oestradiol‐induced tumour growth delay was accompanied by a decrease in the G1 fraction, an accumulation of cells in the S‐phase, and polyploidy, whereas neither treatment with tamoxifen nor ovariectomy influenced cell cycle distribution. The results indicate that oestradiol and tamoxifen have different mechanisms of action. In addition, they were interpreted as indicating different mechanisms regulating ovariandependent tumour growth, on the one hand, and oestrogen and tamoxifen‐induced tumour growth inhibition, on the other. The results support the view that the presence of receptors may be of importance but is not a sufficiently clear marker to allow prediction of the endocrine sensitivity of individual breast tumours.
Infectious transmission of human T‐cell leukemia virus to rabbits
Authors: Miyoshi I. , Yoshimoto S. , Kubonishi I. , Fujishita M. , Ohtsuki Y. , Yamashita M. , Yamato K. , Hirose S. , Taguchi H. , Niiya K. , Kobayashi M.
Published: February 2007

A rabbit lymphoid cell line (Ra‐I) was established by co‐cultivation with a human T‐cell line (MT‐2) carrying human T‐cell leukemia virus (HTLV). The Ra‐l cell line is chromosomally male and is persistently infected with HTLV. Ra‐l cells, with or without mitomycin‐C treatment, were inoculated intravenously (i. v.) into 3 female rabbits. All 3 animals responded with the production of antibodies to HTLV antigens. Lymphocytes from one of these seroconverters were cultured in the presence of T‐cell growth factor (TCGF) and HTLV particles were detected in the TCGF‐grown lymphocytes which were chromosomally female. Co‐cultivation of lymphocytes from the 2 other seroconverters with lymphocytes from 2 anti‐HTLV‐negative healthy men gave rise to the establishment of an HTLV‐producing T‐cell line derived from each individual. Blood transfusion from one of the HTLV‐infected rabbits into 2 female rabbits also resulted in the seroconversion of both recipients. An HTLV‐carrying lymphoid cell line (Ra‐2) was established from one of the transfusion‐related seroconverters. The Ra‐2 cell line was initially TCGF‐dependent but later became TCGF‐independent. These results indicate that HTLV can be transmitted to rabbits. These animals may provide a suitable model system for studying the mode of transmission and pathogenicity of HTLV.
Radioimmunodetection of murine mammary adenocarcinoma (TA3/Ha) lung and liver metastases with radioiodinated PNA
Authors: Shysh A. , Eu S. M. , Noujaim A. A. , Suresh M. R. , Longenecker B. M.
Published: February 2007

Organ‐specific lung and liver metastatic variants of the murine TA3/Ha mammary adenocarcinoma cell line were selected by sequential in vivo growth with intervening in vitro cell culture. These variants readily formed specific lung or liver metastatic lesions upon i. v. injection into A/J mice. TA3/Ha cells produce a large cell surface glycoprotein called epiglycanin, which contains a high proportion of Thomsen‐Friedenreich (TF) antigenic structures. The presence of non‐cryptic TF has been associated with malignancy in humans and animals. We used peanut lectin agglutinin (PNA), which has a preferential affinity for TF antigenic structures, to determine whether these selected metastatic variants retained the TF antigen expression. In vitro, the TA3/Ha etastatic variant lines exhibited strong PNA binding similar to that seen with human RBC after neuraminidase treatment to expose the cryptic TF antigen. In contrast, the non‐epiglycaninproducing TA3/St subline did not bind PNA appreciably. Autoradiography of liver sections with TA3/Ha metastatic lesions after 125I‐PNA i. v. indicated an avid uptake throughout the viable tumor mass and FITC‐PNA staining of these tissue sections readily identified the metastatic tumors under fluorescence microscopy. Tissue biodistribution studies revealed that lung or liver containing the TA3/Ha metastatic variant nodules retained about 7 to 8 times as much of an i. v. dose of radioiodinated PNA as did controls, allowing for clear delineation of tumor‐infiltrated lung or liver by gamma xintigraphy. These in vitro and in vivo tests confirm that the selected organ‐specific TA3/Ha variants retained the binding characteristics of the parent TA3/Ha line. These observations illustrate the potential utility of radiolabelled PNA for the detection of TF‐antigen‐expressing tumors and metastases. This murine system with organ‐specific TA3/Ha metastatic variants also provides a model for evaluation of various other macromolecular probes for tumor radioimmunodetection of metastatic lesions.
Anti‐tumor effect of inelammatory neutrophils: Characteristics of in vivo generation and in vitro tumor cell lysis
Authors: Lichtenstein Alan , Kahle Janet
Published: February 2007

Inflammatory neutrophils elicited by intraperitoneal injection of Corynebacterium parvum, thioglycollate or proteose peptone were capable of lysing different murine and human tumor targets in a shortterm chromium‐release assay. A single‐cell cytotoxicity assay, which evaluated effector‐target cell interactions at the single‐cell level, confirmed a PMN‐mediated tumor‐lytic effect. Optimal lysis was achieved by PMNs obtained 6 hr after injection of C parvurn and 16 hr after injection of thioglycollate. In vitro, loss of tumor cell membrane integrity occurred extremely rapidly following conjugation with inflammatory PMNs (beginning within 15 min of the binding step). By 45 min, the lytic event was completed. Addition of catalase or superoxide dismutase to the cytotoxicity assays prevented tumor lysis in a concentration‐dependent fashion, indicating that hydrogen peroxide and superoxide, products of the PMN respiratory burst, are mediators of the lytic reaction.
An analysis of some risk factors for lung cancer in Hong Kong
Authors: Koo L. C. , Ho J. H‐C. , Lee N.
Published: February 2007

Lung cancer has been the major cause of cancer death in Hong Kong for more than a decade. Although it is known that some 95% of male cases can be attributed to smoking, the etiological factors in women remain elusive. Among “never‐smoked” female cases, increases in attributable risk from passive smoking were limited to only some of the histological types of lung carcinomas, and an overall analysis of all types did not reveal any significant increase in relative risk from this source. Other environmental factors which encourage bronchial irritation are suspected. Methodological differences may explain the differences in proportional distributions of histological lung tumor types noted between previous reports, and the risk values attributed therein to active and passive smoking.
Gastric cancer risk in chronic atrophic gastritis: Statistical calculations of cross‐sectional data
Authors: Slpponen Pentti , Kekki Matti , Haapakoski Jaason , Ihamäki Timo , Siurala Max
Published: February 2007

Relative risk (RR) and cumulative risk of gastric cancer (GCA) were calculated for different grades of atrophic gastritis (AG) of the antrum and body. Cross‐sectional data on the occurrence of AG in a representative population sample (371 subjects), and Finnish Cancer Registry data on GCA were used in the calculations. The RR was increased significantly in severe AG of the antrum and the body (18.1 and 4.6 times, respectively), but not significantly in the less severe grades of AG. As a risk factor, severe antral and body gastritis were independent of each other. The cumulative risk, i. e., the probability of contracting GCA within the following 10 years in age groups 50–54 … 70–74 years was calculated to vary from 2.3% to 9.3% and from 8.7% to 31.9% in severe antral AG and from 0.9% to 4.5% and from 3.6% to 16.6% in severe body AG in males and females, respectively.
Characterization and partial purification of human milk fat globule membrane antigens by polyacrylamide gel electrophoresis and immunoblotting using monoclonal antibodies
Authors: Ashorn Per , Krohn Kai
Published: February 2007

Polyclonal and monoclonal antibodies were used to characterize human milk fat globule (HMFG) membrane antigens, using gel filtration, ion exchange chromatography and western blotting. Although originally generated against HMFG, monoclonal antibodies also reacted with skim milk. In western blotting, several antigen molecules were seen, of which some were detected by all available antibodies, and one only with monoclonal antibody Ill D 5, previously shown to react with mammary and ovarian carcinomas bearing estrogen receptors. This or these antigens, with a molecular weight of about 53 kd, were isolated by fractionation in SDS‐PAGE. A low polypeptide content was demonstrated but the antigenic structure could be stained with periodic‐acid‐Schiff and biotinylated peanut agglutinin, indicating the presence of galactose or N‐acetylgalactosamine residues. While this 53‐kd molecule is the only determinant exclusively stained by III D 5 antibody, it is suggested that it carries epitopes related to estrogen receptor activation.
Transfection by human oncogenes: Concomitant induction of tumorigenicity and tumor‐associated membrane alterations
Authors: Collard J. G. , van Beek W. P. , Janssen J. W. G. , Schijven J. F.
Published: February 2007

NIH 3T3 cells were transfected with DNA derived from human bladder carcinoma, colon carcinoma and HL60 promyelocytic leukemia cells. The transfectants were examined for the presence of human oncogenes in relation to tumorigenic potential and composition of surface‐located fucosyl glycopeptides by gel filtration, Concanavalin‐A‐binding and high‐performance liquid chromatography. All transfectants, harboring 3 different human cellular ras genes, appeared to be tumorigenic in nude mice and displayed characteristically altered glycopeptides. The surface glycopeptides were consistently changed to higher apparent molecular weight due to enrichment in higher‐branched sialic‐acid‐containing glycopeptides. Similar alterations have been found previously in virally‐ and chemically‐transformed cells in vitro and tumors raised in vivo, and were designated as cancer‐related or tumor‐associated glycopeptides. Revertants derived from HL60‐DNA‐induced transfectants, which had lost the transfected human N‐ras oncogene, simultaneously lost their tumorigenic potential and expression of cancer‐related membrane glycopeptides. In addition, spontaneous transformants, exhibiting morphology and growth patterns indistinguishable from those of tumor‐DNA‐induced transfectants, neither contained transferred human DNA sequences nor expressed cancer‐related glycopeptides. Nevertheless these cells were capable, after prolonged latency periods, of inducing tumors in nude mice. Cells derived from such tumors constantly displayed cancer‐related glycopeptides on their surface, suggesting selection of tumorigenic cells from spontaneous transformants during passage in nude mice. In one of these tumors at least, an endogenous mouse ras‐gene appeared to be activated. The results indicate a close correlation between the presence of activated ras‐oncogenes in the genome of the transfected cells, the tumorigenic potential of these cells and the expression of surface‐located cancer‐related glycopeptides. The data suggest that functions provided by human ras‐oncogenes contribute to the alteration of membrane glycopeptides on tumor cells.
Induction of HLA class‐II antigen expression on human carcinoma cell lines by IFN‐gamma
Authors: Schwartz Reinhard , Momburg Frank , Moldenhauer Gerhard , Dörken Bernd , Schirrmacher Volker
Published: February 2007

The effects of recombinant interferon gamma (IFN‐γ) on HLA antigen expression were examined in various human carcinoma cell lines: carcinomas of the lung (ChaGo, Oat 75, SK‐LC‐LL), colon (HT‐29, WiDr), larynx (HEp‐2), cervix (ME‐180) and mammary gland (AIAb). Surface expression of HLA antigens was determined using monoclonal antibodies directed against monomorphic determinants of HLA class II (HD II, 2.06) and HLA class I (W6/32) by several techniques including indirect immunofluorescence, immunoperoxidase assay, radioimmunoassay of cells, and immunoprecipitation of radioiodinated cells. Without previous exposure to IFN‐γ, HLA class‐II antigen was surface‐expressed on a small subpopulation of HEp‐2 only. All cell lines examined, except AIAb, could be induced to cell‐surface expression of HLA class‐II antigen after 48 hr incubation in the presence of IFN‐γ (10 units/ml). HLA class‐I antigen was present on all cell lines even without treatment, and it was increased after IFN‐exposure. Incorporation of 3H‐thymidine and 3H‐leucine and expression of the transferrin receptor were not significantly altered by IFN‐γ application, indicating a specific regulatory effect of IFN‐γ on HLA antigen expression. Our data demonstrate that HLA class‐II surface expression is inducible by IFN‐γ in a variety of epithelial tumor lines. This may have implications for the host immune system defense mechanisms against tumors.
Capacity of B‐lymphocytic lines of diverse tumor origin to produce and respond to B‐cell growth factors: A progression model for B‐cell lymphomagenesis
Authors: Gordon John , Åman Pierre , Rosén Anders , Ernberg Ingemar , Ehlin‐Henriksson Barbro , Klein George
Published: February 2007

Human cell lines established from cases of acute lymphoblastic leukemia, lymphosarcoma, Burkitt's lymphoma and multiple myeloma and representing stages of B‐lymphocyte development ranging from pre‐B through to plasma cells, were assessed for their ability to produce and respond to B‐cell growth factors (BCGF). All B‐cell lines studied were found to be constitutive producers of a growth activity which assisted the S‐phase entry of normal activated B‐cells and provided growth support for lymphoblastoid cells transformed by Epstein‐Barr virus. Furthermore, all lines responded by enhanced proliferation to supernatants from a BCGF‐producing T‐cell hybridoma. Not all lines, however, displayed autostimulation to their own supernatants and no tumor B‐cell line appeared totally dependent on soluble factors for its growth. Non‐tumorigenic B‐cell lines, by contrast, revealed a strict dependency on homologous growith factor for their continued proliferation in suspension culture. The findings support a progression model of lymphomagenesis based upon the utilization, production and, ultimately, emancipation from growth‐promoting soluble factors.
Common acquired naevi and the risk of malignant melanoma
Authors: Green Adèle , Maclennan Robert , Siskind Victor
Published: February 2007

The relationship between common acquired naevi and malignant melanoma of the skin was assessed in a case‐control study. In 183 patients and 183 controls matched for sex, age and area of residence, there was a strong association between the presence of pigmented naevi on the arms and melanoma. In comparison with persons having no naevi on the arms, the crude relative risk of melanoma was 28.0 when naevi were present. After adjusting for hair colour, propensity to sunburn and lifetime sun exposure, the final risk estimate was 30. 1. Family history did not appear to be a determinant of disease independent of the above risk factors.
Seroepidemilogy of human T‐lymphotropic retrovirus type I (HTLV‐I) in residents of niigata prefecture, Japan. Comparative studies by indirect immunofluorescence microscopy and enzyme‐linked immunosorbent assay
Authors: Aoki Tadao , Miyakoshi Hideo , Koide Hiroko , Yoshida Tsutomu , Ishikawa Hiroshi , Sugisaki Yuji , Mizukoshi Mikio , Tamura Kazue , Misawa Hirondo , Hamada Chuya , Ting Robert C. , Robert‐Guroff Marjorie , Gallo Robert C.
Published: February 2007

A large sample of carriers of human T‐lymphotropic retrovirus type 1 (HTLV‐I) in Niigata Prefecture was examined for the detection of natural antibodies to HTLV‐I‐related antigens in sera using both indirect immunofluorescence microscopy (IFM) and enzyme linked immunosorbent assay (ELISA). The present findings are based on multiple surveys, using each assay technique at least twice. Although Niigata Prefecture has been considered a non‐endemic region for HTLV‐I, Sado Island has been proven by this study to be a relatively endemic pocket within this non‐endemic area. Seropositivity was highest in residents of Sado Island; 97/1, 117 (8.7%) by IFM and 33/1, 061 (3.1 %) by ELISA; followed by Niigata City, 18/650 (2.8%) by IFM and 16/638 (2.5%) by ELISA; and lowest in the remaining areas, 57/2, 631 (2.2%) by IFM and 20/2, 551 (0.8%) by ELISA. Seropositivity was demonstrated in 172/4, 398 (3.9%) by IFM and 69/4, 250 (1.6%) by ELISA in Niigata Prefecture taken as a whole. In general, the incidence of seropositive residents increased gradually with age. The sex difference was not significant. The serum samples tested were categorized into 4 groups; (1) IFM and ELISA both positive, (2) IFM positive but ELISA negative, (3) IFM negative but ELISA positive, and (4) IFM and ELISA both negative. By absorption tests, IFM and ELISA seemed to recognize different specific antibodies in sera; IFM recognized antibodies to HTLV‐I‐related cellular antigens in addition to HTLV‐I‐viral antigens, but ELISA recognized antibodies to HTLV‐I viral antigens alone. Thus, IFM detected a broader spectrum of antigens, resulting in recognition of more positive sera than those detected by ELISA.
Biosynthesis of alpha fetoprotein by MCF‐7 human breast cancer cells
Authors: Sarcione Edward J. , Hart Darleen
Published: February 2007

Direct evidence was obtained for de novo synthesis of AFP by MCF‐7 human breast cancer cells per se. Synthesis was demonstrated by L‐14C‐leucine and L‐35S‐methionine incorporation into immunochemically isolated AFP, and confirmed by radioimmunodiffusion and radioimmunoelectrophoresis. This information indicates that AFP synthesis is associated with normal and neoplastic cells of several different histotypes, and suggests that AFP detected and measured previously in primary human breast cancer tissue cytosol (Sarcione et al., 1983) also resulted from in situ biosynthesis by breast cancer cells per se rather than uptake of exogenous AFP originating from extracellular sources. Evidence that AFP obtained after treatment of 14C‐leucine radiolabelled MCF‐7 breast cancer cell protein with 0.4 M KC1 contained 2.6 times more radioactivity than did AFP obtained before such salt treatment is interpreted as indicating that two different molecular species of de novo synthesized AFP existed in breast cancer cells: (1) larger amount of non‐immunoreactive AFP which became immunoreactive and measurable after KC1 treatment, and (2) smaller amounts of free immunoreactive AFP. 14C‐radiolabelled AFP obtained before and after treatment of cell protein with 0.4 M KC1 codiffused, comigrated with alpha, electrophoretic mobility and gave an identical radioimmunologic reaction both with each other and with added carrier human cord serum AFP. Furthermore, preliminary studies indicated that radiolabelled non‐immunoreactive AFP could be separated from lower‐molecular‐weight free AFP by chromatography on Sephadex G‐200. Taken together, these findings suggest that synthesized free AFP was bound as a non‐immunoreactive high‐molecular‐weight macromolecular complex rather than being covalently linked. Our current working hypothesis is that most of the de novo synthesized endogenous AFP in MCF‐7 human breast cancer cells was rapidly and reversibly bound by hydrophobic bonding to a specific cytoplasmic AFP‐receptor.
Effect of cyclosporin‐A (CsA) on the ability of T lymphocyte subsets to inhibit the proliferation of autologous EBV‐transformed B cells
Authors: Bejarano Maria Teresa , Masucci Maria Grazia , Ernberg Ingemar , Klein Eva , Klein George
Published: February 2007

We have analyzed the effect of Cyclosporin‐A (CsA) on the in vitro suppression of Epstein‐Barr virus (EBV)‐induced B‐cell proliferation exerted by autologous T‐lymphocyte subpopulations separated on the basis of cell density. CsA abolished the growth‐inhibitory capacity of high‐density T cells but influenced only marginally the activity of low‐density lymphocytes; this suggested that different mechanisms mediate suppression in the two subsets. Stimulation with irradiated EBV‐transformed cells had a different impact on the activity of low‐ and high‐density lymphocytes. Proliferative and cytotoxic responses were inversely correlated, i.e. high‐density cells proliferated but exerted low cytotoxicity, while the lytic activity of the low density subset was stronger in the absence of significant cell proliferation. Proliferation and generation of cytotoxicity were abrogated by CsA in both subsets. The activities could be restored by addition of exogenous IL‐2, suggesting that the drug may interfere with the cascade of lymphokine–cell interactions which leads to activation of immune responses. From the analysis of the CsA effects on the two subsets it seems that the high‐density one contains specific memory T cells which are activated and proliferate upon encounter with EBV‐infected cells. On the other hand, low‐density lymphocytes are induced to release soluble factors with antiproliferative activity. The secretory function was resistant to the suppressive effect of CsA.
Production of β‐human chorionic gonadotrophin by human squamous carcinoma cell lines
Authors: Cowley Gerard , Smith John A. , Ellison Morag , Gusterson Barry
Published: February 2007

Eight out of 9 human squamous carcinoma cell lines ectopically secrete the beta subunit of human chorionic gonadotrophin (β‐HCG) in amounts detectable by radioimmunoassay. In contrast, both normal and SV40‐transformed keratinocytes show no such evidence of β‐HCG secretion. Ectopic β‐HCG is identical in its gel exclusion and high‐performance liquid chromatographic properties to placental β‐HCG and, in radioimmunoassay, yields a dilution curve parallel to that of placental β‐HCG. Dibutyryl cyclic AMP, but not sodium butyrate, stimulates β‐HCG secretion by the one human cell line examined, LICR‐LON‐HN‐5, suggesting that ectopic production by these cells is similar to trophoblastic β‐HCG secretion, and differs from most other non‐trophoblastic cell lines previously examined.
Immunogenicity of melanoma‐associated gangliosides in cancer patients
Authors: Tai Tadashi , Cahan Leslie D. , Tsuchida Tetsuya , Saxton Romaine E. , Irie Reiko F. , Morton Donald L.
Published: February 2007

The immunogenicity of gangliosides found on human melanoma cells was determined from sera of 26 melanoma patients who were immunized every 1–4 weeks for 4 months with tumor‐cell vaccine (TCV) prepared from cultured melanoma cells. Total lipidbound sialic acid in the gangliosides isolated from TCV was 0.38 μmol/108 cells, and was distributed as follows: 44.8% to GM3, 44.2% to GD3, 5.6% to GM2, and 4.6% to GD2. Sera were tested at monthly intervals for antibodies to each ganglioside by ELISA with purified gangliosides as the antigen source. The immunologic specificity of the antibody was confirmed by absorption tests. None of the 26 patients had detectable anti‐GM3, anti‐GD3, or anti‐GD2 antibodies before immunization, although anti‐GM2 antibody was detected in 3 patients. After immunization, 2 patients developed IgM anti‐GD2, 10 developed IgM anti‐GM2, and 2 developed IgG anti‐GM2 antibodies. No patient developed detectable anti‐GM3 or anti‐GD3 antibodies. These results indicate that both GD2 and GM2 expressed on human melanoma cells are immunogenic in humans, although GM2 appears to be more immunogenic. The other two gangliosides, GM3 and GD3, are present in human sera and in human normal tissues, and thus immunologic tolerance may have been established against these gangliosides. Alternatively, circulating GM3 and GD3 may have neutralized anti‐GM3 and anti‐GD3 antibodies, if any were induced by TCV immunization.
Methylation pattern of human T‐cell leukemia virus in vivo and in vitro: pX and LTR regions are hypomethylated in vivo
Authors: Kitamura Toshio , Takano Masako , Hoshino Hiroo , Shimotohno Kunitada , Shimoyama Masanori , Miwa Masanao , Takaku Fumimaro , Sugimura Takashi
Published: February 2007

The methylation patterns of the gag, pol, env, pX and LTR regions of proviral DNA of human T‐cell leukemia/lymphoma virus type I (HTLV) in fresh leukemic cells and established cell lines were examined using Hpall/Mspl endonuclease. Peripheral blood lymphocytes (PBL) isolated from patients with adult T‐cell leukemia/lymphoma (ATL) did not express viral antigens of HTLV, but PBL that had been cultured for 2 days did express these viral antigens. Most parts of the gag, pol and env regions of the HTLV provirus in PBL isolated from 12 ATL patients and PBL cultured for 2 days were hypermethylated as reported by others. In contrast, in 10 established cell lines that harbored HTLV genomes and expressed viral antigens, HTLV proviruses were hypomethylated. In one cell line, ATL‐IK, which harbored an HTLV genome but did not produce viral antigens, the gag, pol and env regions were hypermethylated. However, two Hpall sites, one in the middle of the gag region and the other in the middle of the pol region, were not methylated even in PBL from most ATL patients. Furthermore, the pX and LTR regions were hypomethylated not only in established cell lines but also in PBL of ATL patients. The hypomethylation of the pX and LTR regions detected in fresh leukemic cells of ATL patients may have some etiological significance in cell transformation by controlling the level of transcription of these regions, or modulating the binding of some factors to these regions.
Sauna, dust and migration as risk factors in lung cancer among smoking and non‐smoking males in Finland
Authors: Tenkanen Leena , Hakulinen Timo , Hakama Matti , Saxén Erkki
Published: February 2007

To study possible causes of the high lung cancer incidence among Finnish males, the Finnish Cancer Registry and the Cancer Registry of Norway in 1962 performed a population survey covering smoking habits, occupational and residential history and, in Finland, sauna habits. The cohort of 4,475 Finnish men has now been followed up for incidence of lung cancer in 1964–1979. The relative importance of 3 factors–sauna baths, occupational exposure to dust, and migration to urban areas–was studied in different smoking categories. Among smokers of ≥ 25 g/day, takers of frequent saunas (more than once a week) displayed some excess risk (RR 1. 7). In smokers an increased risk was also found among those exposed to dust (RR 1.3), and those who had migrated to urban areas (RR 1.8, when compared to native urban population). Dust exposure and migration factors act synergistically with smoking. Migration was the only factor apart from smoking to show a substantial population‐attributable risk, which amounted to 10%.
Adoptive immunotherapy of intracerebral murine lymphomas: Role of different lymphoid populations
Authors: Romani L. , Nardelli B. , Bianchi R. , Puccetti P. , Mage M. , Fioretti M. C.
Published: February 2007

In an attempt to elucidate the cellular mechanisms responsible for the therapeutic activity of systemic immunotherapy in an adoptive transfer system, lymphoma cells were implanted i. c. It was found that, upon peripheral injection of cytotoxic T‐lymphocytes with specificity for the tumor, the cells reached and infiltrated the diseased brain but did not accumulate selectively in the malignant graft. In order to accomplish significant tumor inhibition, the infused lymphocytes, largely expressing the Lyt‐1 +2+ phenotype, apparently cooperated with radioresistant phagocytic cells present in histocompatible hosts and athymic mice.
The relationship between tumour geometry and the response of tumour cells to cytotoxic drugs — An in vitro study using EMT6 multicellular spheroids
Authors: Kwok T. T. , Twentyman P. R.
Published: February 2007

Multicellular spheroids of the EMT6/Ca/VJAC mouse mammary tumour cell line have been used in an investigation of the effect of tumour geometry on the response of tumour cells to 3 cytotoxic drugs, adriamycin (ADM), nitrogen mustard (HN2) and CCNU. In addition to the inherent cellular drug response, factors related to spheroid structure, namely cell‐cycle distribution, intercellular contact, drug penetration and microenvironment (PH, oxygen, glucose, etc.) are believed to influence the response of cells within spheroids to cytotoxic drugs. Selective enzymatic dissociation (with bacterial neutral protease) has been used to separate the cells within large ( ≈ 800 μm in diameter) spheroids into 4 distinct subpopulations. The cells within the subpopulations have been characterized by their DNA content, RNA content, tritiated thymidine labelling in dex, cell size and clonogenic capacity. It was found that cells at the surface of spheroids are relatively larger and more proliferative than cells towards the centre while their clonogenic capacity is similar. Studies on the response of EMT6/Ca/VJAC log and plateau‐phase monolayer cells have been carried out in parallel and have shown that cycling cells are more sensitive to ADM and HN2 than are non‐cycling cells but somewhat less sensitive for the response to CCNU. Since the response patterns of cells from different regions of spheroids to HN2, treated either before disaggregation (intact spheroid) or after disaggregation (isolated spheroid cells), are similar and the surviving fraction increases from the surface towards the centre of the spheroid, cell cycle distribution is thought to be the only factor involved in the cytotoxicity of HN2 towards cells within the spheroids. Although the patterns of response to ADM of cells within intact spheroids and isolated spheroid cells are similar to those for HN2, the initial slope of the curve for intact spheroids is much steeper than that of the isolated spheroid cells. Therefore, in addition to the factor of cell‐cycle distribution, drug penetraction also appears to be involved in the action of ADM on spheroids, while the factors of intercellular contact and microenvironment appear to be relatively less important. The reverse pattern was found for the response of cells within different regions of spheroids to CCNU, treated as intact spheroids or as isolated spheroid cells (i. e., greater killing of inner compared with outer cells). There was, however, a much greater differential effect when treatment was carried out on intact spheroids compared with isolated spheroid cells. As 14C‐CCNU is found to be distributed consistently through out the whole spheroid, both cell‐cycle distribution and changes in the cellular microenvironment within the spheroids appear to play important roles in the response of cells within spheroids to CCNU.
Natural cytotoxicity of human blood monocytes and natural killer cells and their cytotoxic factors: Discriminating effects of actinomycin D
Authors: Uchida Atsushi , Klein Eva
Published: February 2007

The effect of actinomycin D on target susceptibility to human blood natural killer (NK) cells and monocytes was analysed in direct cell‐mediated and their cytotoxic factor‐mediated cytotoxicity assays. Treatment of K562 cells with actinomycin D reduced their susceptibility to lysis by non‐adherent lymphocytes and Percoll‐purified large granular lymphocytes (LGL) in a 4‐hr 51Cr‐release assay, without affecting their sensitivity to monocytes purified by adherence to autologous serum‐coated plastic surfaces. The drug treatment caused no shift in the kinetics of cytotoxicity. In the target binding assay LGL formed fewer conjugates with actinomycin‐D‐treated K562 cells than with untreated ones, while the binding of monocytes to targets was not reduced by the drug treatment of K562 cells. The cold target competition assay revealed that actinomycin‐D‐treated cold K562 cells showed less successful inhibition than untreated cold K562 cells. Lymphocytes and monocytes could be induced to release soluble cytotoxic factors, termed natural killer cytotoxic factors (NKCF) and monocyte cytotoxic factors (MCF), respectively, when co‐cultured with K562 cells. Both cytotoxic factors lysed NK‐sensitive target cells in a 48‐hr assay. Actinomycin‐D‐treated K562 cells reduced or abolished the ability to stimulate the release of NKCF from lymphocytes, whereas they induced MCF secretion from monocytes as effectively as untreated ones. On the other hand, actinomycin D treatment of K562 cells enhanced their susceptibility to NKCF and MCF. This actinomycin‐D‐induced augmentation of target sensitivity to the cytotoxic factors was restricted to NK‐sensitive target cells (K562 and Molt‐4). NK‐resistant target cells (Raji, YAC‐I, EL4 and T blasts) were not lysed by NKCF and MCF even after they were treated with actinomycin D. The capacity of K562 cells to bind NKCF and MCF was not altered by actinomycin D. Treatment of the adherent cell population with OKMI or Leu‐MI plus complement abrogated both cell‐mediated cytotoxicity and MCF production, while Leu‐11b plus complement was ineffective. These results suggest that the effect of actinomycin‐D treatment can be used to distinguish the two distinct types of blood mononuclear cells with natural cytotoxicity, NK cells and monocytes, and that each effector type recognizes different plasma membrane moieties of NK target cells, although the cytotoxic factors released from each effector cell similarly bind to and lyse the target cells.