Growth behavior of low passage cultured cells derived from human non-small cell lung carcinoma in deepithelialized rat tracheas transplanted into nude mice.

Proliferation behavior of six low passage cell lines derived from human non-small cell lung carcinoma was investigated both by cell inoculation into deepithelialized Fisher 344 rat tracheas xenotransplanted into nude mice and by direct cell injection into the subcutis of nude mice. In the tracheal transplants, all the cell lines showed a comparable or higher degree of differentiation than in the primary lesion, while most cell lines showed a lower degree of differentiation in the mouse subcutis. The surfaces of the tracheal transplants repopulated with the cells derived from squamous cell carcinomas were initially lined with a layer of flattened cells, two to three cells thick, while those repopulated with the cells from the three adenocarcinoma cases (HKT-3, HKT-5, HKT-6) were initially lined with simple epithelia comprising cuboidal-, round-and columnar-shaped cells, respectively. Subsequently, the cells from HKT-3 increased in height and proliferated in a pseudostratified pattern, those from HKT-5 proliferated in a papillary growth pattern and those from HKT-6 proliferated in a cribriform pattern. Ultrastructurally, they resembled ciliated bronchial epithelial cells, non-ciliated bronchiolar epithelial cells and bronchial goblet cells, respectively. Histological examination of cultured cells using this system was considered to be advantageous in the evaluation of differentiation since it provided unique proliferation patterns unobserved in primary tumors or tumors formed after s.c. direct cell injections.     

Cellular pharmacokinetics of mercaptopurine in human neoplastic cells and cell lines

The accumulation, metabolism, and retention of mercaptopurine (MP) was studied in four human neoplastic cell lines (three acute leukemia lines Molt-4, CCRF-CEM, and HL-60; and one Burkitt's lymphoma line, Wilson), each of which was sensitive to MP. Two cell lines resistant to MP (WilsonR and CCRF-CEMR) were also studied. The cell lines were incubated for 3 h in 10 microM [14C]MP and then placed in drug-free media for an additional 3 h. Cell samples were obtained at regular intervals, and the intracellular MP metabolites were measured in the acid-soluble fractions by anion-exchange high-pressure liquid chromatography. MP accumulated progressively within cells during the 3-h drug exposure period and declined rapidly when the cells were placed in drug-free media. Over 80% of the intracellular MP was present in the form of three nucleotide metabolites, MP ribose monophosphate, thioxanthosine monophosphate, and thioguanosine monophosphate. MP ribose monophosphate was found in greatest amount, accounting for 59-85% of the intracellular metabolite pool. Thioxanthosine monophosphate thioguanosine monophosphate were detected in lesser amounts. Study of leukemic cells obtained from patients demonstrated a similar pattern of MP accumulation, metabolism, and retention, although the overall amounts of the various metabolites formed were less. In contrast, there was essentially no MP nucleotide metabolite formation in the two MP-resistant cell lines. A more complete understanding of the cellular pharmacokinetics of MP in human neoplastic cells is likely to lead to a more rational use of the drug in the clinical setting.     

Isolation of two syngeneic cell lines from a rat mammary carcinoma: growth factor production by neoplastic epithelial cells

Two distinctly different clonal cell lines were isolated from a mammary tumor induced by ingestion of 7,12-dimethylbenz[a]anthracene (CAS: 57-97-6) in a SD rat. Cells of one of the clones, RMT-1 clone E4, showed typical epithelial characters; it was concluded that they were derived from neoplastic mammary epithelial cells. The other clone, M2, exhibited characters consistent with its derivation from the normal mammary myoepithelial component. The 2 cell lines had different proliferative responses to growth factors (GF) such as cholera toxin, dexamethasone, and epidermal GF. The epithelioid E4 cells were found to produce potent growth-promoting activity in culture medium that stimulated proliferation of myoepithelial M2 cells as well as that of stromal fibroblasts. The present work provides supporting evidence that the mechanism of "paracrine stimulation" is operative in the mammary tumorigenesis of the rat.     

Gap junction expression following UVC-induced neoplastic transformation in human hybrid cell lines

Decreased connexin gene expression and loss of the capacity for either homologous or heterologous intercellular communication has been associated with neoplastic transformation. We tested the hypothesis that loss of gap junctional intercellular communication (GJIC) correlates with tumorigenic potential in the HeLa x skin fibroblast human hybrid cell system. Connexin gene expression, gap junction function and tumorigenicity were determined for the non-tumorigenic somatic hybrid cell line (CGL1) and a series of UVC-induced tumorigenic cell lines derived from CGL1. CGL1 and the parental skin fibroblasts express connexin43 (alpha1 gap junction gene) mRNA and protein, form gap junctional plaques and have functional gap junctions. UVC- irradiation of CGL1 cells produced a cell line (UV12) with an aggressive tumorigenic phenotype, which lost connexin43 expression as well as both homologous and heterologous GJIC and was in this respect similar to HeLa cells. However, the phenotype of UV12 cells exhibited some instability and revertants to a less aggressive tumorigenic phenotype were isolated. These cells expressed connexin43 mRNA and protein, and demonstrated homologous GJIC. Furthermore, cells reconstituted from a tumor derived from this revertant cell line retained significant connexin43 expression and homologous GJIC, although they exhibited an aggressive tumorigenic phenotype. Thus, functional homologous GJIC cannot be dissociated from tumorigenicity in this system. However, heterologous GJIC between these same UVC-induced tumorigenic cell lines and normal human skin fibroblasts was reduced, whereas the non-tumorigenic hybrid cells showed extensive heterologous GJIC. In summary, re-acquisition of connexin43 expression and homologous GJIC does not restore the non-tumorigenic phenotype in UVC- induced tumorigenic HeLa skin fibroblast human hybrid cells. However, reduction of heterologous GJIC does correlate with tumorigenicity in this cell system.      

n-3 and n-6 fatty acid processing and growth effects in neoplastic and non-cancerous human mammary epithelial cell lines.

The type rather than the amount of dietary fat may be more important in breast carcinogenesis. While animal studies support this view, little is known about the effects of essential fatty acids (EFAs) at the cellular level. The MCF-7 breast cancer and the MCF-10A non-cancerous human mammary epithelial cell lines are compared in terms of growth response to EFAs and ability to incorporate and process the EFAs. Eicosapentaenoic (EPA, n-3) and docosahexaenoic (DHA, n-3) acids, presented bound to albumin, inhibited the growth of MCF-7 cells by as much as 50% in a dose-dependent manner (6-30 microM) in medium containing 0.5% serum. alpha-Linolenic (LNA, n-3) and arachidonic (AA, n-6) acids inhibited growth less extensively, while linoleic acid (LA, n-6) had no effect. In contrast, MCF-10A cells were not inhibited by any of the EFAs at levels below 24 microM. The differential effects of AA, EPA and DHA on MCF-7 and MCF-10A cells support a protective role of highly unsaturated essential fatty acids against breast cancer. The EFAs were primarily incorporated into phosphoglycerides. MCF-7 cells showed chain elongations and possibly delta 8 desaturation, but no AA was formed from LA, nor EPA or DHA from LNA. In contrast, MCF-10A cells desaturated and elongated the exogenous EFAs via all the known pathways. These findings suggest defects in the desaturating enzymes of MCF-7 cells. LNA, DHA and AA presented to MCF-7 cells in phospholipid liposomes inhibited growth as extensively as albumin-bound free acids, but were less extensively incorporated, suggesting different mechanisms of inhibition for the two methods.     

Antiproliferative action of isatine-beta-thiocarbohydrazone and N-ethylisatine-beta-thiocarbohydrazone on human PBMC and on two neoplastic cell lines

The antiproliferative action of two synthetic compounds, isatine-b-thiocarbohydrazone (IsTCH) and N-ethyl isatine-beta-thiocarbohydrazone (N-Et-IsTCH), towards healthy human peripheral blood mononuclear cells (PBMC) and two neoplastic cell lines in vitro, was investigated. IsTCH and N-Et-IsTCH were dissolved in DMSO and then diluted with nutrient medium to desired final concentration. Target cells were PBMC, as well as human cervix carcinoma - HeLa cells, and murine melanoma B16 cells. Five different concentrations (3 microM to 50 microM) of investigated agents were applied on target cells. Cell survival was determined 72 h after the agent's action using MTT test. Results obtained showed that both investigated compounds exerted a dose dependent antiproliferative action to neoplastic cell lines. Their action was only cytostatic; trypan blue exclusion test did not show any sign of direct drug cytotoxicity when drugs concentration were less than 50 microM. ICs50 +/- SD for IsTCH antiproliferative action were 61.69 +/- 4.25 microM for HeLa cells; 34.1 +/- 7.15 microM for B16 cells: 17.62 +/- 7.11 microM for nonstimulated and 30.0 +/- 9.46 microM for stimulated (by 5 mg/ml PHA) PBMC. ICs50 +/- SD for the action of N-Et-IsTCH were 21.86 +/- 1.77 microM for HeLa cells; 10.37 +/- 1.55 microM for B16 cells; >47 microM for both, nonstimulated and for stimulated, PBMC. Nonstimulated human PBMC appeared to be the most sensitive to the cytostatic IsTCH action; while HeLa cells were the most resistant. N-Et-IsTCH showed more than two or five fold stronger antiproliferative effect toward B16 cells than on HeLa or PBMC cells, respectively, and more than three times intensive activity compared to IsTCH, indicating specificity of N-Et-IsTCH towards inhibition of melanoma cell growth. While increasing concentrations of IsTCH led to decrease in the the PBMC induced suppression of HeLa cell survival; N-Et-Is-TCH in the difference from IsTCH, in dose dependent way contributed to the PBMC induced suppression of HeLa cell survival. In conclusion, the activity of N-Et-Istch on malignant melanoma cells deserves further investigation.     

Endoglin (CD105) expression in non-neoplastic and neoplastic human tissues and human cancer cell lines

Endoglin is an integral membrane glycoprotein that binds TGF-beta(1,3) with high affinity and is thought to play an important role in modulating the interaction of TGF-beta with its cell surface receptors. In this study a recently characterized monoclonal antibody (29-G8) recognizing endoglin was used to examine expression in a variety of human tissues and human cancer cell lines. Formalin-fixed, paraffin-embedded sections were examined by light microscopy and cell lines were analyzed by now cytometry. Immunostaining was noted in a variety of non-neoplastic epithelia from different organs; most of the neoplastic tissues surveyed also demonstrated prominent immunoreactivity for 29-G8. Flow cytometric analysis of the cell lines revealed strong 29-G8 immunoreactivity in almost all lines examined. Our results suggest that endoglin expression is much more ubiquitous than was previously thought and that endoglin may play a role in modulating TGF-beta binding activity in a variety of normal and neoplastic human tissues.     

Study of the in vitro cytotoxic potential of natural and synthetic coumarin derivatives using human normal and neoplastic skin cell lines

A selection of natural and synthetic coumarin compounds, including the hydroxylated and nitrated derivatives, were assessed for their cytotoxic potential using the microculture 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay for cellular viability. For the first time this study utilized both human skin malignant melanocytes (SK-MEL-31) and normal human skin fibroblastic cells (HS613.SK), allowing identification of those coumarin derivatives that are selectively toxic. Coumarin was found to exhibit comparatively low toxicity in both cell types, while 7-hydroxycoumarin (7-OHC) and coumarin had similar activity in SK-MEL-31 cells. The entire series of hydroxylated coumarins were considerably more toxic in HS613.SK than in SK-MEL-31 cells. Novel synthetic nitrated coumarins, 6-nitro-7-hydroxycoumarin (6-NO2-7-OHC) and 3,6,8-nitro-7-hydroxycoumarin (3,6,8-NO2-7-OHC), were shown to be significantly more toxic to SK-MEL-31 than HS613.SK cells. In the malignant melanocyte skin cell line (SK-MEL-31) the cytotoxic effects of these nitro-derivatives were shown to be dose and time dependent. Therefore, the cytotoxic potential of coumarins appears to be highly dependent on the nature and position of the functional group. In addition, nitration of 7-OHC produced compounds that were cytotoxic to malignant melanocytes, suggesting that these nitro-derivatives may have a chemotherapeutic role in the future.     

Preneoplastic changes of xenotransplanted human distal airway epithelium induced by systemic administration of 4-nitroquinoline-1-oxide to host nude mice

To elucidate a possible role of hematogenously transported carcinogens in pathogenesis of peripheral lung carcinoma in humans, we investigated whether the bronchiolar and alveolar epithelial cells of adult human lung xenografts in nude mice could be a target for the chemical carcinogen 4-nitroquinoline-1-oxide (4NQO) after its systemic administration to the host mice. Peripheral lung tissues from adult humans were transplanted s.c. into nude mice, and 4NQO (15 mg/kg) was administered s.c. to the host mice at a site distant from the xenografts at 2 and 3 weeks after transplantation. The human lung xenografts were maintained for from 20 to 52 weeks, and then serial sections were examined histologically and immunohistochemically. Three types of epithelial changes, i.e. epidermoid metaplasia, papillary hyperplasia of columnar and epidermoid cells, and atypical adenomatous hyperplasia, were induced in the 4NQO group, with a statistically significant difference for these combined epithelial lesions (P less than 0.01) and for epidermoid metaplasia (P less than 0.05) compared to the control group. Some epidermoid metaplasias showed significant nuclear atypia. In addition, almost all foci of epidermoid metaplasia and papillary hyperplasia contained cells positive for carcinoembryonic antigen, suggesting both types of the lesions were preneoplastic. The morphologic characteristics of the atypical adenomatous hyperplasia were very closely similar to those of the hitherto reported preneoplastic or putative neoplastic lesions in the human peripheral lung. Our results indicated that the alveolar and bronchiolar epithelial cells of human lung xenografts were affected by systemically applied 4NQO, and subsequently underwent transformation to a preneoplastic state.     

Characterization of the influence of anti-hormone and/or anti-growth factor neutralizing antibodies on cell clone architecture and the growth of human neoplastic astrocytic cell lines

Summary The influence of five anti-hormone and/or anti-growth factor neutralizing antibodies on thein vitro proliferation of four human astrocytic tumor cell lines (U87, U138, U373, H4) is quantitatively described by means of a new tool which makes it possible to evaluate cell growth and cell clone architecture concomitantly. This tool relies upon the combined use of the digital cell image analyses of Feulgen-stained nuclei and the Delaunay and Voronoi mathematical triangulation and paving techniques. Of the five anti-hormone and/or anti-growth factors tested here, the anti-luteinizing hormone-releasing hormone (LHRH) antibody induced the most marked perturbation in the U138 and U373 cell lines, whereas this role was played by the anti-epidermal growth factor (EGF) antibody in the U87 and H4 cell lines. The anti-gastrin (G) antibody significantly modified the growth and/or cell clone architecture of the U138, U87 and H4 cell lines, as did the anti-transforming growth factor alpha (TGFalpha) antibody. The anti-transforming growth factor beta (TGFbeta) antibody modified the growth and/or cell clone architecture of the four cell lines under study. If the five antibodies are taken into consideration, the results strongly suggest that four (the anti-G, the anti-EGF, the anti-LHRH and the anti-TGFalpha) act as inhibitory agents on some glioma cell line proliferation, while the fifth one, i.e. the anti-TGFbeta, act as a stimulator of cell proliferation, perhaps by abrogating the inhibitory effects of TGFbeta on proliferation. A comparison of cell growth data with cell clone architecture characteristics provided further evidence of some specific influence exercised by a given hormone and/or growth factor on glioma cell proliferation. Indeed, the anti-LHRH antibody caused the most pronounced perturbations in the U138 and U373 cell clone architecture; this feature was observed in the H4 cell line and, to a lesser extent in the U87 one after the anti-EGF antibody had been used.     

Galactosyl transferase activity in human transitional cell carcinoma lines and in benign and neoplastic human bladder epithelium.

Cultured cells of human transitional cell carcinoma line MGH-U1, in suspension, were assayed for galactosyl transferase by measurement of the transfer of [3H]galactose from uridine diphosphate:[3H]galactose to desialylated ovine submaxillary mucin. The assay was optimized with respect to time and to protein, uridine disphosphate:galactose, desialyated ovine submaxillary mucin, and Triton X-100 concentrations. This assay was then applied to fresh specimens of benign, inflamed, and neoplastic bladder epithelium from 33 patients who under went cold-cup biopsies at cytoscopy. Transitional cell carcinoma specimens gave values in the range of 24.7 to 184.8 cpm [3H]galactose transferred per microgram protein per hr [72.0 +/- 44.7 (S.D.); n = 25]; normal and inflamed specimens ranged from 0.8 to 46.1 cpm/microgram protein per hr [8.3 +/- 8.4 (S.D.); n = 35]. By using a known method of cell rupture, cell ghosts, representing cell-surface membranes, were isolated both from the cultured cell line and from two biopsy specimens of transitional cell carcinoma. Although a complete enzymatic and electron microscopic analysis was not undertaken, the coincidence of an enzyme marker with the cell ghost fraction containing the elevated galactosyl transferase made it appear probable that this enzyme is located in the cell surface.     

Analysis of expression of cell surface antigen Mr 74,000 phosphoglycoprotein in normal, oncogene-transformed, and neoplastic rat cell lines

A Mr 74,000 phosphoglycoprotein (gp74) present on the surface of oncogene-transformed murine cells but not untransformed NIH 3T3 cells was previously identified with mouse monoclonal antibody 45-2D9. The original cell population used as the immunogen was found to consist of two cell populations. The purpose of this study was to characterize these cell populations; determine the distribution of gp74 on normal, transformed, and neoplastic cells; and to characterize the gp74 molecule. Southern hybridization studies of cloned cell populations demonstrated that the immunizing cell population consisted of c-Ha-ras-transfected NIH 3T3 cells and Kirsten sarcoma virus-transformed rat cells (TRF cells). TRF cells showed a high level of gp74 expression. We observed that the expression of gp74 was increased on chemically and spontaneously transformed rat cells compared to untransformed rat cells. No binding of monoclonal antibody 45-2D9 was detected to rat adult and fetal tissue. Immunoperoxidase staining, immunofluorescence flow cytometry, and immunoprecipitation analysis of dimethylbenz[a]anthracene-induced metastatic 13762NF rat mammary adenocarcinoma clonal sublines demonstrated an inverse relationship between gp74 expression and metastatic phenotype. gp74 was immunoprecipitated from two low and medium metastatic clonal sublines (MTC and MTF7), but not from highly metastatic clone MTLn3 cells. Biosynthetic labeling and immunoprecipitation studies demonstrated that gp74 was phosphorylated on serine residues and was not secreted from transformed cells. No detectable protein kinase activity in an immune complex assay was associated with this molecule. We conclude that increased gp74 expression by rat cells is associated with transformed and neoplastic cells.     

Neural progenitor cell lines inhibit rat tumor growth in vivo

Current therapies for gliomas often fail to address their infiltrative nature. Conventional treatments leave behind small clusters of neoplastic cells, resulting in eventual tumor recurrence. In the present study, we have evaluated the antitumor activity of neural progenitor cells against gliomas when stereotactically injected into nucleus Caudatus of Fisher rats. We show that the rat neural progenitor cell lines HiB5 and ST14A, from embryonic hippocampus and striatum primordium, respectively, are able to prolong animal survival and, in 25% of the cases, completely inhibit the outgrowth of N29 glioma compared with control animals. Delayed tumor outgrowth was also seen when HiB5 cells were inoculated at the site of tumor growth 1 week after tumor inoculation or when a mixture of tumor cells and HiB5 cells were injected s.c. into Fisher rats. HiB5 cells were additionally coinoculated together with two alternative rat gliomas, N32 and N25. N32 was growth inhibited, but rats inoculated with N25 cells did not show a prolonged survival. To evaluate the possibility of the involvement of the immune system in the tumor outgrowth inhibition, we show that HiB5 cells do not evoke an immune response when injected into Fisher rats. Furthermore, the rat neural progenitor cells produce all transforming growth factor beta isotypes, which could explain the observed immunosuppressive nature of these cells. Hence, some neural progenitor cells have the ability to inhibit tumor outgrowth when implanted into rats. These results indicate the usefulness of neural stem cells as therapeutically effective cells for the treatment of intracranial tumors.     

Oncogene-related growth factors and growth factor receptors in human malignant glioma-derived cell lines

Oncogenes induce malignant transformation of cells. Two oncogenes are closely related to genes coding for a mitogenic growth factor (v-sis to the PDGF gene) and a receptor for a mitogenic growth factor (v-erb B to the EGF receptor gene). We studied the possibility that cells derived from malignant gliomas produce mitogenic factors that bind to cell surface receptors, the activation of which could lead to excessive stimulation of cell proliferation. All six cell lines tested secrete into their medium factors that stimulate DNA synthesis. The factor secreted by one cell line was characterized and found to resemble PDGR Six of 11 cell lines had receptors for PDGF demonstrable by binding and receptor autophosphorylation assays. Six of six cell lines tested had EGF receptors demonstrable by binding and receptor autophosphorylation experiments. The extremely high levels of EGF receptor in one cell line may reflect excessive expression of the erb B oncogene associated with abnormalities of chromosome 7 that occur in this cell line.     

Search for specific markers of neoplastic epithelial duct and myoepithelial cell lines established from human salivary gland and characterization of their growth in vitro

The neoplastic epithelial duct cells human salivary gland (HSG) and myoepithelial cells human pleomorphic adenoma (HPA) established from human salivary gland were examined by the immunoperoxidase method for the presence of specific antigens such as carcinoembryonic antigen (CEA), S-100 protein, secretory component (SC), lactoferrin (LF), and myosin. Isolation of the cells and their morphologic features were reported previously. Consequently, the presence of CEA, SC, and LF in the HSG cells was demonstrated. The HPA cells were identified to express the specific antigens reactive to anti-S-100 protein, anti-myosin and anti-CEA sera in addition to the presence of oxytocin receptor. When the two cell lines were co-cultured in monolayer culture or within the sponge matrix, a large number of ductlike or tubular structures were formed in an optimal ratio of 1:2 in HSG and HPA cells, whereas the cultures of HSG cells only grew with occasional formation of ductlike structure. In addition, in HSG and HPA cells in an area with their contact in the mixed cultures, CEA staining was intensified as compared with the culture of HSG or HPA cells only and further S-100 protein was detected in HSG cells, whereas S-100 protein was not detected in the culture of HSG cells only. These findings strongly suggest that the intercalated duct and myoepithelial cells from human salivary gland propagate with their interaction together in the expression of specific antigens such as CEA and S-100 protein or in the morphogenesis of salivary gland epithelial cells.     

All-trans-retinoic acid inhibits the growth of human rhabdomyosarcoma cell lines

We have been evaluating the role of all-trans-retinoic acid (RA) in the differentiation and growth of human rhabdomyosarcoma (RMS) cell lines. Treatment of both embryonal (RD) and alveolar (RH30) human RMS cell lines with all-trans-RA resulted in a dose-dependent inhibition of cell growth with a maximal inhibition of 92 and 66%, respectively, at 5 x 10(-6) M. When 13-cis-RA was used under identical experimental conditions, maximal growth inhibition was 41 and 37%, respectively. This stereo-specific growth inhibition was not associated with morphological or biochemical evidence of myogenic differentiation. Furthermore, all-trans-RA demonstrated no evidence of competition with binding of insulin-like growth factor II (IGF-II), an autocrine growth factor in RMS, to its membrane receptor as evaluated by an [125I]IGF-I-receptor-binding assay. Attempts to rescue all-trans-RA growth-inhibited RMS cells with exogenous IGF-II resulted in no increase in growth compared to cells treated with all-trans-RA alone. We conclude that RA inhibits the growth of human RMS cell lines in a dose-dependent, stereo-specific manner, is not associated with differentiation, and does not appear to be directly related to IGF-II.     

Production of transforming growth factor-alpha in human tumour cell lines

Forty-one human tumour cell lines were examined for the production of epidermal growth factor (EGF)/transforming growth factor (TGF)-alpha-like activity (EGF/TGF-alpha-LA), immunoreactive (IR-) EGF and IR-TGF-alpha. EGF/TGF-alpha-LA was determined by radioreceptor assay, in which the factors with capacity to bind to EGF receptor could be detected. IR-EGF and IR-TGF-alpha were determined by the respective radioimmunoassays. Both EGF/TGF-alpha-LA and IR-TGF-alpha were detected in 11 tumour cell lines. The levels of EGF/TGF-alpha-LA correlated well with those of IR-TGF-alpha. A small amount of IR-TGF-alpha was detected in five other lines. In contrast, IR-EGF was not detectable in any of the 41. Consequently, it can be concluded that EGF/TGF-alpha-LA produced by human tumour cells is mainly TGF-alpha rather than EGF. It was also revealed that melanoma cell lines produce a large amount of TGF-alpha frequently. Gel filtration studies revealed that TGF-alpha produced by melanoma cell lines was identical to human (h) TGF-alpha(1-50), except for one line, in which IR-TGF-alpha with a different molecular size was detected. Northern blot analysis revealed that bands corresponding to hTGF-alpha mRNA were present in melanoma cell lines producing a large amount of IR-TGF-alpha, indicating that the TGF-alpha produced is the product of hTGF-alpha gene. Further studies are required to discover the actual biological roles of TGF-alpha produced by melanoma cells as well as other types of cancer cells.     

Transforming growth factor-alpha secretion by epidermal growth factor-dependent human tumor cell lines

Pairs of cell lines from spontaneous human tumors (cervical adenocarcinoma, melanoma, and synovial sarcoma) were established using serum-free culture conditions with and without exogenous epidermal growth factor (EGF). EGF-adapted cultures of melanoma and cervical adenocarcinoma origin secreted higher levels of bioactive transforming growth factor alpha (TGF-alpha) when compared to cultures maintained in the absence of EGF. Depletion of EGF for these EGF-adapted cultures resulted in growth arrest. In contrast, the sarcoma cell lines did not secrete TGF-alpha regardless of the culture conditions but EGF significantly stimulated proliferation of these cells in short-term assays. We show that exogenous EGF induces TGF-alpha production and supports proliferation of tumor cells of various tissue origin but is not essential for in vitro growth factor-deprived conditions.     

Interferon potentiates cytotoxic effects of 5-fluorouracil on cell proliferation of established human cell lines originating from neoplastic tissues

A potentiation of the cytotoxic effects of 5-fluorouracil (5-FU) on human tumor cells by interferon was examined. The human neoplastic cell lines used were HeLa (uterine cervical cancer), MCF-7 (mammary cancer), WI-38 CT-1 (embryonic lung fibroblasts transformed in culture by Co-60 gamma-ray irradiation), KMM-1 (myeloma) and Raji (Burkitt's lymphoma). The normal human cell strain used was WI-38 (normal human lung fibroblasts). The cytotoxic effects were determined by colony formation for HeLa, MCF-7, WI-38 CT-1 and WI-38 cells, and by cell growth for KMM-1 and Raji cells. Each cell line was different in sensitivity to interferon or 5-FU. Interferon potentiated synergistically the cytotoxic effects of 5-FU on HeLa, WI-38 CT-1 and KMM-1 cells. In the case of Raji cells, the cytotoxic effects of the combination of interferon and 5-FU were additive. Neither synergistic nor additive lethal effects of the combination of the 2 agents were observed in MCF-7 and WI-38 cells. The present results indicate a possibility that interferon and 5-FU can mutally reduce the amount of the other needed to treat cancer patients.