bluuebunny/crossref_metadata_2025 huggingface dataset:
Estrogen suppression of erbB2 expression is associated with increased growth rate of ZR‐75‐1 human breast cancer cells in vitro and in nude mice
Authors: Wärri Anni M. , Laine Aire M. , Majasuo Karin E. , Alitalo Kari K. , Härkönen Pirkko L.
Published: February 2007

Amplification and enhanced expression of the erbB2/HER‐2/neu gene has been associated with an increased growth rate and poor prognosis of human breast cancer. We have studied the relationship between erbB2 expression and the regulation of cell growth by estrogen and anti‐estrogens in the human breast cancer cell line ZR‐75‐1 in vitro and in athymic nude mice, pS2 being used as a marker gene for estrogen‐stimulated gene expression. Only low amounts of erbB2 mRNA were seen in the cells grown in vitro in the presence of estrogen which stimulated the cells to proliferate rapidly and induced the expression of pS2 mRNA. Upon hormone withdrawal, erbB2 mRNA and protein increased, while pS2 mRNA declined to an undetectable level and cell proliferation slowed down. Opposite but more rapid changes were observed upon estrogen addition. The anti‐estrogens toremifene and tamoxifen inhibited estrogen induction of pS2 expression, down‐regulation of erbB2 expression and proliferation of the ZR‐75‐1 cells in a concentration‐ dependent manner. Similar results were obtained in nude mice. ZR‐75‐1 cells formed tumors only in mice carrying estrogen pellets. In these tumors little erbB2 mRNA was seen. Concomitant administration of toremifene or tamoxifen increased erbB2 mRNA and abolished pS2 mRNA. Our results show that enhanced expression of erbB2 is associated with hormone deprivation and growth arrest of the estrogen‐dependent breast cancer cell line ZR‐75‐1. Thus, in mammary epithelial cells, erbB2 may have important estrogen‐regulated functions which are not related to cell proliferation.
Human T‐cell recognition of epstein‐barr virus‐induced replication antigen complexes
Authors: Pothen Susan , Richert John R. , Pearson Gary R.
Published: February 2007

Cellular immune responses to Epstein‐Barr Virus (EBV)‐associated antigens play an important role in the control of EBV‐immortalized B lymphocytes. The nature of the antigens that serve as targets for these responses remains largely unknown. The purpose of the experiments reported here was to determine if virus‐replication‐associated antigens might function as targets for T‐cell immunity with the emphasis on EBV‐induced early antigen (EA) complex. Activated T‐cell populations directed against this group of antigens would theoretically be effective in controlling the production of new virus progeny. Our results demonstrate that polypeptides associated with the EA complex do, in fact, induce the proliferation of memory T‐cells from EBV‐infected individuals irrespective of their serological status to the EA complex. The major polypeptide associated with the diffuse component of EA (EA‐D) was notably effective in inducing a strong proliferative T‐cell response. Cell lines established from EBV‐infected individuals following continuous exposure to either p17 or p50 components of the EA complex over a 10‐ to 12‐week period were composed primarily of CD4‐positive T‐cells, although CD8‐positive cells also persisted for up to 9 weeks in culture. The data suggest that components of the EA complex might function as important target antigens in the immunosurveillance of EBV‐infected cells.
Contribution of glutathione and glutathione‐dependent enzymes in the reversal of adriamycin resistance in colon carcinoma cell lines
Authors: Lai Gi‐Ming , Moscow Jeffrey A. , Alvarez Manuel G. , Fojo Antonio T. , Bates Susan E.
Published: February 2007

Four human colon cancer cell lines (SW620, LS 180, DLD‐I, and HCT‐15) and sub‐lines isolated in vitro by selection with Adriamycin were studied for reversal of intrinsic and acquired Adriamycin resistance, using buthionine sulfoximine (BSO) to deplete cellular glutathione alone and in combination with the P‐glycoprotein antagonist verapamil. GSH levels varied among the parental cell lines but did not increase with resistance. In the parental SW620, DLD‐1 and HCT‐15 and their drugresistant derivatives, there was no relation between the effect of the glutathione‐depleting agent BSO, the mRNA expression of both selenium‐dependent glutathione peroxidase (GPx) and glutathione S‐transferase pi (GSTπ), bulk glutathione S‐trans‐ferase (GST) activity, and the degree of resistance. However, in LS 180 and its derivative sub‐lines, which do not principally rely on P‐glycoprotein (Pgp) for Adriamycin resistance, treatment with BSO demonstrated a relatively diminished GSH depletion and enhanced recovery. In comparison with the other acquired cell lines, BSO specifically reversed acquired resistance in the LS 180 Adriamycin‐resistant subline (LS 180 Ad 150) after short‐term drug exposure. Furthermore, the LS 180 Ad 150 cells demonstrated an increase in both GPx and GSTπ mRNA expression. These observations suggest that glutathione‐mediated detoxification of Adriamycin may play a role in the resistance of this sub‐line. Verapamil enhanced Adriamycin cytotoxicity 1.2‐ to 12‐fold in the intrinsically resistant cells and as much as 15‐fold in cell lines with acquired resistance. Combination of BSO with verapamil resulted in additive, but not synergistic, reversal of resistance. The results underscore the complex nature of Adriamycin resistance, and suggest a role for drug‐resistance‐modulating agents in the treatment of colon carcinoma.
Interferon‐gamma interrupts autocrine growth mediated by endogenous interleukin‐6 in renal‐cell carcinoma
Authors: Gruss H.‐J. , Brach M. A. , Mertelsmann R. H. , Herrmann F.
Published: February 2007

Cells of the renal‐cell carcinoma line ACHN constitutively produce IL‐6, express the IL‐6 receptor and use IL‐6 in an autocrine fashion to augment their growth. Growth arrest of ACHN cells can be achieved by exposure of cells to neutralizing monoclonal antibody (MAb) to recombinant human (rh) IL‐6. Treatment of ACHN cells with rh IFN‐γ also leads to inhibition of proliferation of these cells in a dose‐dependent manner, that can be reversed by exogenous rh IL‐6, while IFN‐α, IL‐2, IL‐4 and vinblastine or 17‐β‐estradiol has no effect on growth (3H‐thymidine uptake) of ACHN cells and IL‐6 expression. Studies on the mechanism of action of IFN‐γ revealed that IFN‐γ was acting by inhibiting the autocrine IL‐6‐mediated loop generated by ACHN.
Cell proliferation and vascularization in human breast carcinomas
Authors: Monschke F. , Müller W.‐U. , Winkler U. , Streffer C.
Published: February 2007

Cell proliferation and vascularization were studied in 10 human breast carcinomas by an immunoenzyme technique. The monoclonal antibody (MAb) Ki‐67 was used as a marker for proliferating cells and a polyclonal antibody directed against human von‐Willebrand factor to identify blood vessels. The proportion of Ki‐67‐labelled cells varied from 1% to 20%, the number of small blood vessels from 4.4/mm2 to 57.6/mm2. Within single histological sections of individual tumours the percentage of proliferating cells was not related to the number of small blood vessels. However, after evaluation of 5 sections of each tumour, the average values showed that tumours with high grade of vascularization had a higher percentage of Ki‐67‐positive cells than poorly vascularized samples. The influence of vascular density on cell proliferation was investigated in a selected area of one of the tumours (in 2‐dimensions) and with regard to the over‐ and underlying sections (in 3‐dimensions). After 2‐dimensional evaluation, distances from proliferating cells to the closest blood vessel between 10 and 390 μm were observed, and after 3‐dimensional evaluation none of the proliferating cells measured was located more than 130 μm away from the closest vessel.
Human allogeneic melanoma‐reactive T‐helper lymphocyte clones: Functional analysis of lymphocyte‐melanoma interactions
Authors: Radrizzani M. , Benedetti B. , Castelli C. , Longo A. , Ferrara G. B. , Herlyn M. , Parmiani G. , Fossati G.
Published: February 2007

Lymphocyte clones were isolated from CD4+ peripheral‐blood lymphocytes (PBL) of melanoma (Me) patient 9923 (HLA‐DR7, DQw2, w6), co‐cultured for 30 days with autologous accessory cells, allogeneic Me (Me 1811) (HLA‐DR7, DQwl, w2), IL‐1 β (2 U/ml) and IL‐2 (15 IU/ml). The 55 clones tested displayed a CD3+, CD4+, CD8−, T‐cell receptor (TCR) cu/p+, y/8‐ phenotype. Twenty clones were assayed for proliferation the presence of Me 1811 and B‐lymphoblastoid cell line (LCL) 1811, both expressing HLA‐class‐1 and ‐11 (DR7 and DQw2 shared with patient 9923), intercellular adhesion molecule‐I (ICAM‐ 1) and lymphocyte‐function‐associated antigen‐3 (LFA‐3) molecules. Eight clones were found t o be reactive to Me 1811 but not to LCL 1811. Specificity analysis of these 8 clones revealed that each of them proliferated only to Me 1811, not to other 14 Me and 12 different LCL, suggesting recognition of melanoma‐associated antigen (MAA) expressed on the stimulating Me. One clone (103) was analyzed in more detail. A wider specificity analysis showed that it reacted to Me 1811 but not to 10 other Me expressing or not HLA‐DR7, 5 normal melanocyte cultures (2 of them typing HLA‐DR7‐positive when exposed to interferon‐γ ‐ IFN‐γ), 4 tumors other than Me and 20 different LCL. Clones did not show proliferation in the presence of autologous Me cells. Clone proliferation in response to Me 1811 was significantly inhibited by monoclonal antibodies (MAbs) directed to CD3, TCR α/β, TCR β chain V12, CD4 and HLA‐DR. Moreover, following stimulation with Me 1811, clone 103 showed increased surface expression of CD25 (IL‐2 receptor) and CD71 (transferrin receptor) and produced significant amounts of IL‐2 and IFN‐γ. The supernatant taken from co‐culture of clone 103 with Me 1811 augmented the cytotoxicity of PBL 9923 and other allogeneic PBL against K562 and Me 1811. Thus, the lymphocyte clone 103 is a CD4+ Th clone which uses its CD3/TCR α/β complex to recognize an MAA in conjunction with HLA‐DR7. Availability of this type of reagent may prove useful to identify and characterize MAA recognized by lymphocytes.
Cell kinetic analysis of human brain tumors by in situ double labelling with bromodeoxyuridine and iododeoxyuridine
Authors: Hoshino Takao , Ito Satoyuki , Asai Akio , Shibuya Makoto , Prado Michael D. , Dodson Barbara A. , Davis Richard L. , Wilson Charles B.
Published: February 2007

Fifty‐seven patients with brain tumors (29 gliomas, 23 meningiomas, 5 miscellaneous) received infusions of intravenous iododeoxyuridine (IUdR) and bromodeoxyuridine (BUdR) 1–5 hr apart shortly before tumor removal. Excised tumor specimens were stained sequentially for BUdR and IUdR. The percentage of BUdR‐labelled cells was determined to establish the labelling index (LI), or S‐phase fraction, and the ratio of cells labelled only with IUdR to cells labelled with BUdR or with BUdR and lUdR was determined to calculate the duration of S‐phase (Ts) and the potential doubling time (Tp) of each tumor. The BUdR LIs varied from < 1% to 20%, reflecting the malignancy of each tumor. Despite the difference in LIs, however, Ts was fairly uniform (mean ± so, 8.7 ± 2.0 hr). Tp varied from 2 days to more than 1 month and correlated closely with the BUdR LIs (Tp = 23/LI0.93; r2 = 0.91). Double‐labelling studies with IUdR and BUdR allow the S‐phase fraction, Ts and Tp to be determined from a single biopsy specimen and thus provide more useful information on the growth characteristics of individual tumors than can be obtained by single‐labelling studies with BUdR.
Lack of association between in vitro clonogenic growth of human cervical carcinoma and tumour stage, differentiation, patient age, host cell infiltration or patient survival
Authors: Davidson S. E. , West C. M. L. , Hunter R. D.
Published: February 2007

Biopsies from 117 patients with cervical carcinoma were studied using a clonogenic assay to assess in vitro growth. Successful colony growth was achieved in 84 tumours (72%) with a mean colony‐forming efficiency (CFE), based on total viable nucleated cell counts, of 0.18 ± 0.49% (± 1 standard deviation). There was a wide range of values, from 0.003–4.28%, with a coefficient of variation of 272%. The relationship between clinical features of cervical carcinoma and in vitro colony formation was investigated. No significant association was demonstrated between in vitro growth and either clinical stage (r = 0.02), tumour differentiation (r = −0.08) or patient age (r = −0.12). There was no significant difference in tumour grade between the group of tumours which failed to grow in culture and those which grew well (p = 0.70). In addition, there was no correlation between CFE and the degree of macrophage (r = 0.001), lymphocyte (r = 0.12), or granulocyte (r = 0.08) infiltration. There was no significant difference between CFEs of tumours from patients who had died and from those who were alive and well after a minimum of 2 years' follow‐up after radiotherapy (p = 0.51). Ability to form colonies in agar was not associated with a worse prognosis (p = 0.49). Although CFE is an independent biological parameter, our results suggest that, for cervical carcinoma, it is not useful as a univariate prognostic factor.
Apparent fusion of basement membranes in colorectal carcinoma
Authors: Hewitt Robert E. , Powe Desmond G. , Holland Cathrine M. , Gray Trevor , Turner David R.
Published: February 2007

Previous studies on colorectal carcinomas indicate that consistent differences in epithelial basement membrane (EBM) integrity are present between the tumour centre and periphery. We report that within the tumour centre, EBM staining between back‐to‐back (BTB) neoplastic glands (i.e., adjacent glands in direct contact with no intervening connective tissue) generally follows a pattern different from that of EBM staining at the tumour:stromal interface (TSI). Such distinctions are important, since the factors responsible for EBM deficiencies may vary with intra‐tumoural location, as may the prognostic significance of these deficiencies. Analysis of paraffin sections from 130 colorectal carcinoma cases showed that EBM staining between BTB glands is generally weaker and more discontinuous than at the TSI, sometimes appearing as a linear array of immunostained granules on high‐resolution light microscopy. By double‐labelling immunofluorescence analysis of cryostat sections from 30 cases, a decrease in type‐IV collagen:laminin staining intensity ratio was found between BTB glands. Hence, the composition of EBM between BTB glands appears to be abnormal. As much recent evidence indicates that epithelial:mesenchymal interactions play an essential role in EBM formation, the demonstration of immunostained EBM fragments between BTB glands requires an explanation: We suggest that the synthesis of EBM between BTB glands involved previously intervening stromal (mesenchymal) cells, and that EBM fusion and dissolution occur between BTB glands following the displacement of these cells.
Haemopoietic cell kinetics in humans treated with rGM‐CSF
Authors: Lord Brian I. , Gurney Howard , Chang James , Thatcher Nicholas , Crowther Derek , Dexter T. Michael
Published: February 2007

We have investigated the kinetics of myeloid cell proliferation in the marrow of patients with small‐cell lung cancer and treated with 10 daily subcutaneous injections of granulocyte/macrophage colony‐stimulating factor (GM‐CSF). Bone marrow, obtained before and during treatment with the growth factor, was labelled in vitro with tritiated thymidine (3H‐TdR). A 3rd bone‐marrow sample was obtained I hr following an intravenous injection of 3H‐TdR. Subsequent daily blood samples were also collected, and 3H‐TdR labelling was assessed on these and the marrow preparations by autoradiography. GMCSF treatment increased the peripheral granulocytic cells nearly 5‐fold, but this included significant eosinophilia, so that the neutrophilic granulocytes increased only 3.3‐fold. These cells were released from the marrow over a normal time scale, but their peripheral half‐life was about 6 times longer than normal and they were probably functionally defective. Furthermore, significant numbers of immature cells were released from the marrow. Neutrophil production stimulated by GM‐CSF was thus overestimated by measurement of the apparent peripheral granulocytosis. Increased labelling indices and grain counts in the proliferating granulocytic cells of the marrow indicate shortened cell‐cycle times, and the excess granulocyte production appears to be the result of extra amplification divisions in the proliferative compartments.
Stefins and lysosomal cathepsins B, L and D in human breast carcinoma
Authors: Lah Tamara T. , Kokalj‐Kunovar Majda , Štrukelj Borut , Pungerčar Jože , Barlič‐Maganja Darja , Drobnič‐Košorok Marina , Kastelic Lili , Babnik Joža , Golouh Rastko , Turk Vito
Published: February 2007

In the study of 50 matched pairs of breast carcinoma and normal breast tissue, the activities of cysteine proteinases (CPs), cathepsin (Cat) B and Cat L in tumors were increased on average by 18.5‐fold and 52.5‐fold respectively. The differences in activity of cysteine proteinase inhibitors (CPIs) between tumor and control breast tissues was also observed: in approximately two thirds of carcinomas, lowered CPI activity was measured (group‐I patients), while similar or higher tumor CPI activity was measured in the remaining samples (group‐II patients). Relative increases in specific activity of Cat B and Cat L in group I were significantly higher than in group II. In group I more patients with histopathological tumor grade III and negative estrogen (ER) and progesteron receptor (PR) levels were found, but the metastatic involvement of regional lymph nodes was similar in both groups. A 2‐year follow‐up study showed a significant inverse correlation between disease‐free survival and increased Cat L activity, but the differences in group I and group II patients were not significant in this short time interval. In 20 matched pairs of breast carcinoma and normal breast tissue, the mean activity of Cat D was 5.8‐fold higher in tumors compared with controls. The hypothesis that elevated Cat D activity increased CP activity and/or lowered tumor CPI activity due to post‐translational proteolytic modification appeared less likely, since no correlations between corresponding activities were observed. We suggested that lowered CPI might rather reflect changes in transcription of intracellular CPIs, the stefins. Immunoassay and Northern blot analysis showed that the average value of stefin A protein and mRNA content respectively in the majority of investigated breast carcinoma samples were lowered, suggesting the possible value of stefin A in diagnosis and/or prognosis of the disease.
Cancer control programme in india: Opportunities for implementation and evaluation
Authors: Sankaranarayanan R. , Nair M. Krishnan , Mathew Babu , Wesley Remani , Mayadevi S.
Published: February 2007

Cancer is now being appreciated in India as an emerging public health problem. Approximately 600,000 new cancer cases occur in India every year, and the absolute number of new cancer patients will increase considerably, due to growth in the size of the population and an increase in the proportion of elderly persons due to improved life expectancy following control of communicable diseases. This emerging problem has received the attention of the Government of India, and some state governments within the Indian Federation have formulated National and State Cancer Control Programmes to deal with the situation. In the event of these programmes being implemented, consideration should be given now to measures of evaluation of the activities, as many of the indices, used to monitor programmes in developed countries, such as 15% reduction from peak cancer mortality, or in peak mortality from specific cancers, are not applicable in developing countries like India. Factors such as the pattern of tobacco habits in the community, knowledge, attitude and practice (KAP) patterns regarding cancer in the general population, referral practices, and the national pattern of extent of disease at presentation, gain considerable importance from the evaluation point of view. Our article mainly deals with the sources and the quality of baseline data for such factors and the realistic quantitative goals which could be set for the above factors.
MCF‐7 breast cancer cells grown as multicellular spheroids in vitro: Effect of 17β‐estradiol
Authors: Olea Nicolás , Villalobos Mercedes , De Almodóvar J. Mariano Ruiz , Pedraza Vicente
Published: February 2007

To obtain multicellular spheroids from MCF‐7 human breast cancer cells we adhered to the following procedure: (a) limiting the adherence of cell to the substratum; (b) seeding more than the minimum number of cells; (c) guaranteeing the presence of estrogens in the culture medium. Charcoal‐dextran (CO)‐ treated sera seemed to inhibit spheroid formation. A reduction in the concentration of CD‐human sera (from 10% to 5%) added to phenol‐red‐free medium facilitated progress from cellular aggregates to multicellular spheroids. Once the spheroids became initiated, size increased at a rate that showed a good fit to a Gompertzian equation (A = 0.368 ± 0.067 α = 0.065 ± 0.013, r range = 0.890–0.989). Three different patterns of spheroid morphology and proliferative kinetic were defined: (a) spheroids with diameter < 200 μm had a constant pattern of heterogeneity in the distribution of 3H‐TdR‐labelled cells and in the expression of estrogen receptors; (b) spheroids 250 to 700 μm in diameter showed a decrease in the proportion of 3H‐TdR‐labelled cells accompanying inward progression (50% in the outer shell, less than 10% in a cell layer located at a depth of 150 μm) while, at a depth of 170 μm, of signs of concurrent cellular degeneration and death were apparent; and (c) spheroids with a diameter of > 750 μm showed a crust of viable cells uniformly labelled with thymidine without impairment of the proportion of labelled cells when progressing inward from the spheroid crust. The larger the spheroid volume, the lower its growth fraction and the longer its volume doubling time. The hormone‐dependence of MCF‐7 cells in forming multicellular spheroids represents a unique experimental model for assessing estrogen action on cell organization and proliferation.
Detection of human papillomavirus dna sequences in tongue squamous‐cell carcinoma utilizing the polymerase chain reaction method
Authors: Shindoh Masanobu , Sawada Yukiharu , Kohgo Takao , Amemiya Akira , Fujinaga Kei
Published: February 2007

Twenty‐four cases of tongue squamous‐cell carcinoma (SCC) were analyzed for human papillomavirus (HPV) DNAs by the polymerase chain reaction (PCR) method and the dot‐blot hybridization technique. HPV DNAs were detected in 8 cases. One specimen histopathologically diagnosed as poorly differentiated grade‐III SCC contained both HPV‐16 and HPV‐18 DNA, and 7 other cases contained HPV‐16 DNA.
Localized formation of micronuclei in the oral mucosa and tobacco‐specific nitrosamines in the saliva of “reverse” smokers, khaini‐tobacco chewers and gudakhu users
Authors: Stich Hans F. , Parida Bibhuti B. , Brunnemann Klaus D.
Published: February 2007

“Reverse”‐cigar smokers (who hold the burning end of cigars within the mouth), dippers (who place a mixture of Khaini‐tobacco and slaked lime into the lower gingival groove) and users of tobacco‐containing toothpaste (gudakhu) in Orissa, India, were examined for precancerous oral lesions, the frequency of micronucleated cells at 3 different intra‐oral sites, and levels of tobacco‐specific nitrosamines (TSNA) in the saliva. Among reverse‐cigar smokers, a high incidence of leukokeratosis nicotina palati, an elevated frequency of micronucleated cells in the palate (2.5% as compared to 0.6% in non‐smokers and non‐chewers of tobacco) and tongue (2.1 %) from which carcinomas preferentially develop, and up to 5890 ppb nitrosonomicotine and up to 1880 ppb N‐nitrosoanatabine in the saliva were found. Among Khaini‐tobacco chewers, the frequency of micronucleated cells was elevated to 2.1 % in the gingival groove, and up to 1580 ng N‐nitrosonornicotine, 690 ng N‐nitrosoanatabine, 90 ng N‐nitrosoanabasine, and 180 ng 4‐(methyl‐N‐nitrosamino)‐I‐(3‐pyridyl)‐1‐butanone per ml of saliva were observed. The localized elevation of the frequency of micronuclei and cancer development is probably due to a synergistic effect of hyperthermia and tobacco‐related carcinogens among reverse‐cigar smokers, and to the close, prolonged contact between the mucosa and tobacco among Khaini‐tobacco/slaked lime dippers. Neither pre‐cancerous lesions nor an elevated frequency of micronuclei were seen in the oral mucosa of users of gudakhu, a tobacco‐containing toothpaste, which may be due to the low amount of TSNA released from the gudakhu and the short exposure time, which is restricted to the period of tooth brushing.
Laboratory determination of chemotherapeutic drug resistance in tumor cells from patients with leukemia, using a fluorometric microculture cytotoxicity assay (FMCA)
Authors: Larsson Rolf , Kristensen Jörgen , Sandberg Charlotta , Nygren Peter
Published: February 2007

An automated fluorometric microculture cytotoxicity assay (FMCA) based on the measurement of fluorescence generated from cellular hydrolysis of fluorescein diacetate (FDA) to fluorescein was employed for chemotherapeutic‐drug‐sensitivity testing of tumor‐cell suspensions from patients with leukemia. Fluorescence was linearly related to cell number, and reproducible measurements of drug sensitivity could be performed using fresh or cryopreserved leukemia cells. A marked heterogeneity with respect to chemotherapeutic drug sensitivity was observed for a panel of cytotoxic drugs tested in 43 samples from 35 patients with treated or untreated acute and chronic leukemia. For samples obtained from patients with chronic lymphocytic and acute myelocytic leukemia, sensitivity profiles for standard drugs corresponded to known clinical activity and the assay detected primary and acquired drug resistance. Individual in vitro/in vivo correlations indicated high specificity with respect to the identification of drug resistance. The results suggest that the FMCA may be a simple and rapid method for in vivo‐representative determinations of chemotherapeutic drug resistance in tumor cells obtained from patients with leukemia.
Effects of sandostatin and castration on pancreatic carcinogenesis in rats and hamsters
Authors: Meuers M. , Woutersen R. A. , van Garderen‐Hoetmer A. , Barker G. H. , de Jong F. H. , Foekens J. A. , Klijn J. G. M.
Published: February 2007

The effects of treatment with the somatostatin analogue Sandostatin, separately and in combination with surgical castration, on the development of azaserine‐induced lesions in rat pancreas and N‐nitrosobis(2‐oxopropyl)amine (BOP)‐induced lesions in hamster pancreas were investigated. The animals were divided in 4 groups and treated as follows: (a) controls, injected s.c. with saline solution (0.9% NaCI); (b) orchiectomy directly after the last treatment with carcinogen; (c) Sandostatin (SMS 201–995) subcutaneously; (d) orchiectomy followed by treatment with Sandostatin. No significant suppressive effects on plasma EGF or IGF‐I concentrations were noted after Sandostatin treatment, but plasma gastrin levels decreased slightly in the rats, not in the hamsters. In rats, Sandostatin treatment enhanced rather than inhibited growth of acidophilic atypical acinar cell nodules. In hamster pancreas, by contrast, Sandostatin inhibited the development of putative pre‐neoplastic ductular lesions. There was no interaction between treatment with Sandostatin and surgical castration. It was concluded that Sandostatin, when administered prophylactically, has an inhibitory effect on the growth of putative pre‐neoplastic ductular, but not acinar, lesions.
Establishment and characterization of a thymic carcinoma cell line (Ty‐82) carrying t(15;19)(ql5;p13) chromosome abnormality
Authors: Kuzume T. , Kubonishi I. , Takeuchi S. , Takeuchi T. , Iwata J. , Sonobe H. , Ohtsuki Y. , Miyoshi I.
Published: February 2007

A new human cell line, designated Ty‐82, was established from the pleural effusion of a 22‐year‐old woman with undifferentiated thymic carcinoma. This cell line consisted of primitive cells that were positive for α‐naphthyl butyrate esterase and acid phosphatase. The cells were shown to express epithelial membrane antigen, but were completely negative for cytokeratin, carcinoembryonic antigen, glial fibrillary acidic protein, desmin, S‐100 protein, lysozyme, Leu‐7, HLA‐DR (la), leukocyte common antigen, Ki‐I antigen, T‐cell antigens, B‐cell antigens, myelomonocyte antigens, and Epstein‐Barr‐virus nuclear antigen. Electron microscopy showed that the cells were highly anaplastic, with no sign of cellular differentiation to any lineages. The Ty‐82 cell line was found to have a karyotype of 46, XX, t( 15; 19)(q 15;p 13), being identical to that of the patient's tumor cells. Four of 5 nude mice inoculated sub‐cutaneously with Ty‐82 cells developed tumors which displayed a histological picture similar to the original tumor. Thymic carcinoma is a recently recognized entity, and its cellular and clinical behavior are poorly understood. The newly established thymic carcinoma cell line would provide a useful tool for the better understanding of this rare disease.
Biphasic effect of camp‐elevating agents on ICAM‐1 expression stimulated by retinoic acid and interferonγ
Authors: Bouillon Marlène , Fortier Michel A. , Boulianne Robert , Audette Marie
Published: February 2007

Incubation of the human glioma cell line HS 683 in the presence of IFN‐7 or retinoic acid strongly stimulates the cell‐surface expression of the intercellular adhesion molecule ICAM‐I. We have investigated the role of the cAMP‐mediated signal transduction pathway in this process and report that pharmacological agents which increased the intracellular levels of cAMP exhibited a biphasic action on ICAM‐1 expression in human glioma cell line HS 683. Treatment for I hr with 25 (μM forskolin or 1 mM) isobutylmethylxanthine, or for 12 hrwith 100 ng/ml pertussis toxin or 50 fig/ml cholera toxin transiently stimulated ICAM‐1 expression with a maximal level of expression 8 hr post treatment, after which time ICAM‐1 expression returned to the basal level. On the other hand, such pre‐ treatments inhibited the inducing effects of either retinoic acid or IFN‐γ. Indeed, 24 hr after treatment with cAMP‐elevating agents, both the retinoic‐acid‐ and the IFN–γ‐induced ICAM‐1 expression were inhibited by 60 to 80%, with a maximal 90 to 100% inhibition 72 hr post treatment. This inhibition of the cell‐surface expression of ICAM‐1 was confirmed at the mRNA level. The intracytoplasmic levels of cAMP were also quantified following treatments with forskolin, retinoic acid or IFN–γ. In response to forskolin, cAMP levels increased 30‐fold within 5 min, whereas a 10‐fold increase occurred 60 min following treatment with 10 μM retinoic acid. lnterferonγ, in contrast, did not induce cAMP accumulation. These results were also correlated with an in vitro activation of adenylyl cyclase activity by retinoic acid and inhibition of this activity by IFN‐γ, in a dose‐dependent and a GTP‐dependent manner. Our results suggest that the suppression of IFN‐γ‐induced ICAM‐1 expression, obtained upon pre‐treatment with cAMP‐elevating agents, is due to direct antagonism with IFN‐γ action on adenylyl cyclase. However, the inhibition of retinoic‐acid‐induced ICAM‐1 expression cannot be explained by the same mechanisms. The timing of adenylyl cyclase stimulation and cAMP accumulation, as well as the levels of cAMP accumulation, are probably involved in this inhibition. Our results also emphasize the fact that the induction of ICAM‐1 expression is a multi‐step process implicating different transductional signals among which cAMP might be involved as a second messenger.
The relationship between multi‐drug resistance and resistance to natural‐killer‐cell and lymphokine‐activated killer‐cell lysis in human leukemic cell lines
Authors: Treichel Robin S. , Olken Sarah
Published: February 2007

Results concerning a possible link between susceptibility to natural‐cell‐mediated immune cytolysis and the multi‐drug resistance (MDR) phenotype are conflicting. We evaluated in human acute lymphocytic leukemia the relationship between acquired drug resistance and susceptibility to cytolysis mediated by endogenous, interferon‐activated, and interleukin‐2‐ activated natural cytotoxic celts. Eight human leukemia drug‐ resistant/sensitive cell line pairs were evaluated; drug‐resistant sub‐lines included those selected for primary resistance to adriamycin, etoposide, teniposide, vincristine, and vinblastine. A majority of P‐glycoprotein‐positive MDR sub‐lines displayed slight but statistically significant resistance to endogenous and/or interferon‐activated natural‐killer(NK)‐cell‐mediated lysis, as compared with the drug‐sensitive parental type. P‐glycoprotein‐ negative sub‐lines displayed variable NK susceptibility; within this group, the variants selected for primary etoposide resistance were more susceptible to NK cytolysis than parental cells. Results of cold‐target‐inhibition experiments suggest that altered NK susceptibility does not arise solely from modulation of NK target recognition and adherence structures. IL2‐activated killer (LAK) cells lysed both drug‐sensitive and drug‐resistant lines. Two MDR lines selected for primary etoposide resistance displayed enhanced LAK susceptibility. In contrast, the 2 variants selected for resistance to adriamycin exhibited partial resistance to LAK‐mediated killing, which could be overcome at high effector‐to‐target ratios. Our results support the development of interleukin‐2/LAK immunotherapy for the treatment of leukemias with acquired drug resistance.
Isolation and characterization of a monoclonal antibody, K1, reactive with ovarian cancers and normal mesothelium
Authors: Chang Kai , Pastan Ira , Willingham Mark C.
Published: February 2007

We have isolated a new monoclonal antibody (MAb), K1, that reacts with an epitope on the surface of human ovarian carcinoma cells. This antibody was generated by immunization of mice with periodate‐treated human ovarian carcinoma (OVCAR‐3) cells. These mice had been previously made tolerant with normal human kidney membranes. Spleen lymphocytes from these mice were selected prior to fusion using a panning purification method on living OVCAR‐3 cells. Initial screening of surface‐reactive clones was performed in a single day using immunofluorescence on living OVCAR‐3 cells, and secondary screening was performed using immunoperoxidase histochemistry on cryostat sections of normal human tissues and human tumors. The K1 clone was subcloned and identified as an IgM isotype, but was subsequently isotype‐switched to lgGIK using a panning selection method. When evaluated by immunohistochemistry, the antigen reactive with K1 was found in many ovarian non‐mucinous tumors, as well as in squamous tumors of the esophagus, and cervical cancer. The only normal adult human tissues showing uniform reactivity with K1 were the mesothelia of the peritoneal, pleural and pericardial cavities. There was also limited reactivity with epithelia of the trachea, tonsil and Fallopian tube. A similar tissue reactivity for K1 was found in tissues from cynomolgus monkeys. K1 reacted with many of the same tissues and tumors as the previously identified antibody OC125, but several lines of evidence indicate that K1 reacts with a different epitope and probably a different molecule, when compared to OC125. This evidence included assays employing immunofluorescence competition, double‐ label immunofluorescence, and solid‐phase and live‐cell radioim‐ munoassays. Since our data indicate that the antigen reactive with the K1 antibody is a new molecular species, we have named the antigen CAKI. Unlike the shed antigen CA125, CAKI was only cell‐associated and was not found in the supernatant of cultured OVCAR‐3 cells or in the blood of ovarian cancer patients. The K1 antibody may be useful as a targeting agent for therapy and in the diagnosis of ovarian carcinoma, as well as some other human cancers.
Allelic loss on chromosome 22 correlates with histopathological predictors of recurrence of meningiomas
Authors: Sanson Marc , Richard Stéphane , Delattre Olivier , Poliwka Marc , Mikol Jacqueline , Philippon Jacques , Thomas Gilles
Published: February 2007

Meningiomas are common tumors of the nervous system. Although usually benign, they may exhibit variable degrees of aggressiveness. Their probability of recurrence after subtotal resection has been correlated with several histological parameters. Independently, a loss of chromosome 22, as evidenced either by cytogenetics or by somatic loss of alleles, has been observed in about half of the cases studied. In 34 meningiomas we have examined the relationship between loss of chromosome 22 alleles and 6 histological predictors of recurrence. Significant correlations were found for 3 of these, i.e. prominent nucleoli (p < 0.002), microscope count of mitoses (p < 0.05) and nuclear pleomorphism (p < 0.02). Correlation with the other 3, i.e. sheeting of cells, vascularity and micronecrosis, did not reach significance. Total tumor score, defined by the sum of the individual scores for these 6 parameters, was strongly correlated to allelic loss (p < 0.0001). Thus, the loss of chromosome 22 alleles, which possibly contribute to the inactivation of tumor‐suppressor gene(s), might be a potent genetic marker of the aggressiveness of meningiomas.
Different fatty‐acid profiles in phosphoinositides from human fibroblastic meningiomas with or without chromosome 22 monosomy
Authors: Bordoni Alessandra , Hrelia Silvana , Biagi Pier Luigi , Berra Bruno
Published: February 2007

The fatty‐acid composition of phosphoinositides derived from 13 human fibroblastic meningiomas, divided according to the presence or the absence of monosomy of chromosome 22, was analyzed. Phosphoinositides were separated into their 3 main components: phosphatidylinositol, phosphatidylinositol‐4‐phosphate, and phosphatidylinositol‐4,5‐bisphosphate. In all the tumors, regardless of cytogenetic differences, remarkable differences were detected, between phosphatidylinositol and phosphatidylinositol‐4‐phosphate or phosphatidylinositol‐4,5‐bisphosphate. In meningiomas with 22 monosomy, there was a significant decrease in the relative amount of arachidonate in all the phosphoinositide fractions; this situation might determine diminished down‐regulation of the corresponding diacylglycerol, derived by the phospholipase‐C‐stimulated hydrolysis of phosphoinositides, and, consequently, different protein‐kinase‐C stimulation.
Human T‐cell leukemia virus type I(HTLV‐I) infection of T cells bearing T‐cell receptor γδ: Effects of HTLV‐I infection on cytotoxicity
Authors: Yasukawa Masaki , Inatsuki Akira , Yakushijin Yoshihiro , Kobayashi Yuzuru
Published: February 2007

In order to clarify the cellular tropism of human T‐cell leukemia virus type I (HTLV‐I) and the effects of HTLV‐I infection on T‐cell functions, we investigated the infectiousness of HTLV‐I on T cells bearing T‐cell receptor (TCR) γδ and functional alterations of the HTLV‐I‐infected TCR‐γδ+ T cells. CD3+CD4−CD8−TCR‐γδ+ T‐cell clones which possessed cytotoxicity were co‐cultured with a HTLV‐I‐producing T‐cell line. After several weeks, integration of HTLV‐I proviral DNA in TCR‐γδ+ T cells was detected by Southern blot analysis. During the continuous culture of HTLV‐I‐infected TCR‐γδ+ T‐cell clones, 2 distinct phases were observed in terms of cytotoxic activity and expression of the CD3‐TCR‐γδ complex. Early after HTLV‐I infection, TCR‐γδ+ T cells lost their spontaneous cytotoxicity, but this was restored by the addition of lectin. At this time, no differences were observed in the expression of various surface molecules between HTLV‐I‐infected and uninfected parent cells, except for increased expression of CD25 on HTLV‐I‐ infected cells. At about 30 weeks after HTLV‐I infection, the cytotoxicity of HTLV‐I‐infected cells was almost completely lost, even in the presence of lectin, and expression of the CD3‐TCR‐γδ complex on the cell surface was markedly decreased. Concomitant with the decreased expression of CD3‐ TCR‐γδ complexes, a decrease in the elevation of cytoplasmic Ca2+ concentration induced by anti‐CD3 and anti‐TCR monoclonal antibodies (MAbs) was also observed. Our present findings thus show that HTLV‐I can infect TCR‐γδ+ T cells, and that consequently their functions are profoundly affected through 2 distinct phases.
Increased phosphorylation of nuclear phosphoproteins in human lung‐cancer cells resistant to cis‐diamminedichloroplatinum(II)
Authors: Nishio Kazuto , Sugimoto Yoshikazu , Kasahara Kazuo , Fujiwara Yasuhiro , Nishiwaki Shinji , Fujiki Hirota , Ohata Masahiro , Saijo Nagahiro
Published: February 2007

A novel non‐phorbol‐ester‐like tumor promoter, okadaic acid (OA) has been shown to be an inhibitor of protein phosphatase I and IIA and, thus, to cause an “apparent activation” of protein kinase C (PKC). We previously showed that cis‐diamminedichloroplatinum(II) (CDDP)‐resistant cells, PC‐9/CDDP, were cross‐ resistant to OA and that the cross‐resistance was not due to the increased efflux of OA. We hypothesized that the phosphorylation status of some cellular proteins might be important in CDDP‐resistancc. No significant difference in PKC activity or total protein phosphatase activity measured in vitro was seen between PC‐9 and PC‐9/CDDP cells, nor in their sensitivity to inhibition by OA, nor in the amount of phosphorylation of whole cells or TCA‐insoluble material. By SDS‐PAGE after incubation of intact cells with 32P, we detected a marked increase, compared to PC‐9 cells, in phosphorylation of the nuclear proteins of MW 32 and 20 kDa in CDDP‐resistant PC‐9/CDDP cells with no apparent difference in protein content. When phosphorylation of nuclear proteins observed in PC‐9/CDDP cells was analyzed by 2‐dimensional SDS‐PAGE, the 32‐kDa protein had a PI of about 4.5. The 32‐kDa and 20‐kDa bands were increased in a dose‐dependent manner by CDDP treatment. On the other hand, no increase in phosphorylation of these proteins was observed in parental PC‐9 cells. These results demonstrate a marked difference in the phosphorylation status of specific nuclear proteins between parental and CDDP‐resistant cell lines, which may be related to CDDP‐resistance.
Interleukin‐1 produced by tumorigenic fibroblasts influences tumor rejection
Authors: Zöller Margot , Doudevani Amos , Segal Shraga , Apte Ron N.
Published: February 2007

Oncogene‐transformed BALB/c‐3T3 fibroblasts which spontaneously or upon immune‐activation with cytokines and lipopolysaccharide (LPS) generate IL‐I α, were tested for their tumorigenicity as well as their interaction with natural immune defense by NK cells and macrophages. Oncogene‐transformed fibroblasts were weakly tumorigenic, since not all mice developed tumors despite application of high doses of tumor cells. This was independent of the immune status of the host. However, in the immunocompetent host those transformed fibroblast lines which spontaneously produced IL‐I α grew only transiently and then regressed. After induction of IL‐I α production, a decrease in the rate of tumor take was noted and the rate of regression of developing tumors was increased. Regression of IL‐I‐producing transformed fibroblasts was strongly reduced but not completely abolished in sublethally irradiated mice. This indicated that IL‐I production may predominantly influence T‐cell‐mediated defense, but some influence on non‐ adaptive immunity could not be excluded a priori. IL‐I production did not influence susceptibility of transformed fibroblasts towards NK cells and macrophages. However, IL‐I‐producing transformed fibroblasts were most potent stimulators of NK cells and macrophages, the stimulatory effect being locally restricted. In conclusion, IL‐I producing, oncogene‐transformed fibroblasts which generated the cytokine constitutively or upon immune‐activation, were rejected from the tumor‐bearing host following initial growth. Fibroblast‐induced local activation of NK cells and macrophages was shown to play some role in tumor graft rejection. The influence of IL‐I production of transformed fibroblasts on T‐cell‐mediated defense is addressed in the accompanying report.
Characterization of epithelial phenotypes in mortal and immortal human breast cells
Authors: Paine Terry Maloney , Soule Herbert D. , Pauley Robert J. , Dawson Peter J.
Published: February 2007

We have previously described the mortal human breast epithelial culture MCF‐10M, that was derived from fibrocystic breast tissue, was cultivated in medium with low calcium content for over 2 years, and spontaneously gave rise to the immortal MCF‐10 cell line. The emergence of immortalized cells, characterized by growth in conventional calcium levels, from mortal cells has proven to be a reproducible event. Here we report the establishment of a second immortal line from MCF‐10M, designated MCF‐10‐2, and establishment of the MCF‐12 immortal line after long‐term cultivation of MCF‐12M mortal cells from reduction mammoplasty tissue. DNA fingerprinting demonstrated the independent, human origin and lineage of the MCF‐10‐2 and MCF‐12 cell lines. Both lines require cortisol and EGF for maximal growth. The expression in these cultures of in vivo breast epithelial phenotypes was analyzed using 2‐dimensional gel Western blots and immunoper‐ oxidase staining with antibodies to cytokeratins and polymorphic epithelial mucin. MCF‐10M and MCF‐12M retain the cytokeratin profile of the luminal cell (7, 8, 18, 19), and also express cytokeratin 14, found predominantly in basal cells. The immortal lines express a similar profile, except that cytokeratin 19, a component of the fully differentiated luminal cell, is not expressed in the more uniform population seen in MCF‐10 and MCF‐12, but is retained in the morphologically mixed, less‐ selected population of MCF‐10‐2. Epitopes on the polymorphic epithelial mucin, recognized by antibodies HMFG 1, HMFG 2 and SM‐3, were detected in the mortal cultures and in the immortal lines, indicating the occurrence of both normal and abnormal mucin processing. MCF‐10, MCF‐10‐2 and MCF‐12 cells do not form tumors in nude mice, but appear to organize as duct‐like structures before regressing in the 5th week post injection.
Induction of IL‐4 secretion by the radiation leukemia virus (RadLV): Role in autocrine growth stimulation of RadLV infected pre‐leukemic cells
Authors: Yefenof Eitan , Ela Catherine , Kotler Moshe , Vitetta Ellen S.
Published: February 2007

Intrathymic inoculation of radiation‐leukemia virus (RadLV) into C57BL/6 mice induces a population of pre‐leukemic (PL) T cells which progress into clonal, mature thymic lymphomas after a latency period of 3 to 5 months. In order to understand how PL cells are retained in the thymus for a prolonged period of time we determined whether RadLV infected cells secrete and/or respond to a T‐cell growth factor that may be involved in the long‐term maintenance of a thymic PL‐cell pool. We have previously found that in vitro proliferation of RadLV‐infected PL cells is IL‐4‐dependent. Here we show that RadLV induces IL‐4 secretion and IL‐4 receptor (IL‐4R) expression in normal thymic lymphocytes. RadLV‐infected PL thymocytes express IL‐4R and secrete IL‐4. Their IL‐4 secretion could be enhanced if incubated in the presence of RadLV and this enhancement was inhibited by anti‐RadLV antibodies. Several RadLV‐induced lymphoma lines secreted IL‐4 and/or expressed IL‐4R, but these features were not essential for their continuous growth. The results suggest that RadLV induces IL‐4‐dependent autocrine growth which maintains a population of PL T cells in the thymus. Transition from a PL state to overt thymic lymphoma involves emancipation of a PL cell from IL‐4 dependency.