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Objectives: This study aimed to determine whether drug-coated balloon (DCB) angioplasty following intraplaque wiring and the use of modified balloons is safe and effective in the percutaneous treatment of coronary chronic total occlusions (CTOs)., Background: DCB is an alternative therapeutic option without the limitations of permanent vascular implants. However, its efficacy in CTOs has yet to be confirmed. The combination of modified balloons and DCB can be effectively applied when the intraplaque passage of the guidewire is achieved in CTOs., Methods: Data from 124 consecutive CTO lesions (105 patients) treated at our hospital between February 2016 and December 2020 were screened for inclusion and retrospectively analyzed. Among the 118 lesions successfully recanalized, intraplaque wiring was achieved in 108, and 85 were treated by the DCB-only approach following cutting/scoring balloon dilatation., Results: Follow-up data were available for 82 lesions (71 patients). The median occlusion length was 18.5 mm, and the J-CTO score was 1.7 ± 0.9. No in-hospital major adverse cardiac events occurred, including abrupt vessel closure. During the median 29-month follow-up period, target lesion revascularization was performed for 10 lesions. Follow-up coronary angiography (8.7 ± 3.9 months after the index procedure) was performed for 64 lesions, demonstrating late lumen loss of -0.15 mm (interquartile range -0.4 to 0.23 mm), binary restenosis (diameter stenosis ≥50%) in 12 lesions (18.8%), and late lumen enlargement in 37 (57.8%)., Conclusion: The DCB-only approach following the use of modified balloons is a promising strategy for coronary CTOs when intraplaque wiring is achieved., (© 2023 Wiley Periodicals LLC.)

Objectives: This study aimed to examine whether the combination of excimer laser coronary atherectomy (ELCA) and drug-coated balloon (DCB) angioplasty can provide feasible clinical outcome in patients with ST-segment elevation myocardial infarction (STEMI) with 8-month and 2-year scheduled follow-up angiography., Background: Intracoronary thrombus elevates the risk of interventional treatment in patients with STEMI and hampers drug absorption into the vasculature released from DCB., Methods: Sixty-two patients with STEMI within 24 h after the onset of symptoms were enrolled in this prospective, single-center, single-arm study., Results: The laser catheter was successfully crossed distal to the culprit lesion in all cases. No ELCA-related adverse events occurred. Bail-out stenting was required in two patients (3.2%) after adjunctive ballooning; thus, the remaining 60 patients were completed with DCB angioplasty without stenting. Scheduled angiography at 8 months and 2 years was completed in 100% and 85.2%, respectively, and minimal lumen diameters were 3.4 ± 0.5, 3.4 ± 0.6, and 3.4 ± 0.5 mm after the procedure, at 8 months and at 2 years, respectively. Binary restenosis was observed in five patients (8.1%) in whom target lesion revascularization was performed. The duration of dual antiplatelet therapy was 2.3 ± 2.2 months, and neither abrupt vessel closure, reinfarction, cardiac death nor major bleeding was observed., Conclusion: A combination of DCB angioplasty with ELCA is a feasible therapeutic option for STEMI., (© 2022 Wiley Periodicals LLC.)

Objectives: To evaluate the feasibility and safety of a virtual 3-Fr system [5-Fr sheathless-guiding catheter (GC)] for percutaneous coronary intervention (PCI)., Background: The use of miniaturized devices for PCI is gaining popularity because of increased patient comfort and decreased risk of access site complications., Methods: From July 2010 to December 2012, consecutive patients who underwent elective PCI (planned or ad hoc PCI) at our hospital were enrolled. PCI using the virtual 3-Fr system was attempted as our initial strategy, unless a 6-Fr or larger GC was considered to be suitable [lesions with heavy calcification, large (>2 mm) side branches, or chronic total occlusion]., Results: Five hundred sixty-six patients underwent elective PCI during the study period, and 132 patients who met the criteria underwent PCI using the virtual 3-Fr system. Procedures using the virtual 3-Fr system were successful in 126 patients (95%); 111 (84%) were performed using the transradial approach, 110 (83%) were ad hoc procedures, and 45 (31%) were complex coronary lesions (type B2 or C). Six patients required conversion to a conventional 5- or 6-Fr sheath and catheter system. No intraoperative complications occurred, and radial artery patency was achieved in all patients who underwent transradial procedures., Conclusions: PCI using the virtual 3-Fr system is a feasible and viable alternative to conventional procedures that use a sheath and GC in appropriately selected patients. This small-caliber system may minimize endovascular trauma, particularly during transradial coronary procedures., (© 2014 Wiley Periodicals, Inc.)

Kaposi's sarcoma-associated herpesvirus (KSHV), the etiologic agent of Kaposi's sarcoma, causes malignancies frequently in patients with acquired immunodeficiency syndrome. In the United States and Europe, KSHV infection is common among men who have sex with men. However, the seroprevalence of KSHV among men who have sex with men in Japan is unknown. In the present study, the seroprevalence of KSHV was investigated among 230 men who have sex with men and 400 age- and area of residence-matched men (controls) using a mixed-antigen (KSHV-encoded K8.1, open reading frame 59, 65, and 73 proteins) enzyme-linked immunosorbent assay and an immunofluorescence assay. Among the Japanese men who have sex with men, serological assays revealed that 27 (11.7%) were seropositive for KSHV; 20 (5%) of the men in the control group were also KSHV seropositive. The seroprevalence of KSHV among men who have sex with men was significantly higher than in the control group (odds ratio = 2.52, 95% confidence intervals = 1.38-4.62, P = 0.0019, Chi-square test). Infection with the human immunodeficiency virus, Treponema pallidum, or hepatitis B and C virus did not correlate with KSHV infection. Furthermore, the association of KSHV seropositivity with specific sexual activities was not statistically significant. In conclusion, a higher KSHV seroprevalence was found among Japanese men who have sex with men than among the controls, suggesting that the circulation of KSHV infection is more efficient among men who have sex with men in Japan than among men who do not engage in such sexual activities., (Copyright © 2013 Wiley Periodicals, Inc.)

The 2009 pandemic H1N1 influenza A virus spread quickly worldwide in 2009. Since most of the fatal cases were reported in developing countries, rapid and accurate diagnosis methods that are usable in poorly equipped laboratories are necessary. In this study, a mobile detection system for the 2009 H1N1 influenza A virus was developed using a reverse-transcriptase loop-mediated isothermal amplification (RT-LAMP) kit with a disposable pocket-warmer as a heating device (designated as pwRT-LAMP). The pwRT-LAMP can detect as few as 100 copies of the virus--which is nearly as sensitive as real-time reverse-transcription polymerase chain reaction (RT-PCR)--and does not cross-react with RNA of seasonal influenza viruses. To evaluate the usefulness of the pwRT-LAMP system, nasal swab samples were collected from 56 patients with flu-like symptoms and were tested. Real-time RT-PCR confirmed that the 2009 H1N1 influenza A virus was present in 27 of the 56 samples. Of these 27 positive samples, QuickVue Influenza A+B immunochromatography detected the virus in only 11 samples (11/27; 40.7%), whereas the pwRT-LAMP system detected the virus in 26 of the 56 samples (26/27 of the positive samples; 96.3%). These findings indicate that the mobile pwRT-LAMP system is an accurate diagnostic system for the 2009 H1N1 influenza A virus, and has great potential utility in diagnosing future influenza pandemics., (Copyright © 2011 Wiley-Liss, Inc.)

Angiogenesis is an essential process for tumor growth and is regulated by tumor-derived angiogenic cytokines. Osteopontin (OPN) is one of the cytokines produced by various tumor cells and is suggested to be involved in angiogenesis by upregulating endothelial cell migration in cooperation with vascular endothelial cell growth factor (VEGF). To provide evidence of OPN involvement in a causal role in tumor angiogenesis, we generated a stable transfectant from murine neuroblastoma C1300 cells to constitutively secrete high levels of murine OPN. The OPN mRNA expression and protein secretion were confirmed by RT-PCR and ELISA, respectively. The biological activity of secreted OPN was determined with migration assay by using human umbilical vein endothelial cells (HUVEC). Transfection with OPN gene did not increase VEGF production and did not affect gene expression of other angiogenic factors confirmed by complementary DNA macroarray system. To demonstrate the effect of OPN on tumor-induced angiogenesis in vivo, millipore chambers containing OPN-transfected or control cells were implanted to the dorsal air sac of mice. The OPN-transfected cells significantly induced neovascularization in comparison to the control cells in mice. Conclusively, these results provide direct evidence of OPN involvement in the role of tumor angiogenesis., (Copyright 2002 Wiley-Liss, Inc.)

Selective tyrosine kinase inhibitors are regarded as promising antitumor agents for cancer treatment. Iressa (ZD1839) is an orally active, selective EGFR-TKI (epidermal growth factor receptor-tyrosine kinase inhibitor) that blocks signal transduction pathways implicated in cancer cell proliferation, survival and other host-dependent processes promoting cancer growth. The cellular mechanisms of ZD1839 action against human malignant cells and drug-resistant cells were evaluated in vitro. Among the cell lines tested, ZD1839 showed a strong growth-inhibitory effect in vitro on human leukemic cells resistant to phorbol ester. This cell line, K562/TPA, shows a non-P-glycoprotein-mediated multidrug-resistant phenotype. The IC50 value of ZD1839 on K562/TPA was approximately 400-fold lower than that on the parental K562 cell (K562 = 12 +/- 2 microM; K562/TPA = 0.025 +/- 0.002 microM) in vitro as determined by a dye formation assay. The expression of EGFR and EGFR mRNA was clearly present in K562/TPA but not in parental K562 cells as determined by Western blotting and RT-PCR. EGFR was autophosphorylated in K562/TPA detected by the antiphosphotyrosine antibody. The in vivo antitumor effects of ZD1839 on K562 and K562/TPA cells were also investigated in BALB/c nude mice. K562/TPA cells transplanted subcutaneously into mice disappeared completely with ZD1839 treatment (20 mg/kg/day, days 3-9). This was not the case in K562 cells. These results suggest that ZD1839 is highly active against tumor cells with non-P-glycoprotein-mediated multidrug resistance that express EGFR. Iressa is a trademark of AstraZeneca (Cheshire, UK)., (Copyright 2001 Wiley‐Liss, Inc.)

Aragusterol A (YTA0040), isolated from the Okinawan marine sponge of the genus Xestospongia, is a potent anti-tumor marine steroid that possesses a unique structural component. This compound showed broad-spectrum anti-proliferative activity against a panel of 14 human cancer cell lines (IC(50) = 0.01-1.6 microM). P-glycoprotein-mediated, multidrug-resistant cells showed cross-resistance to YTA0040 cells, whereas cisplatin-resistant non-small-cell lung-cancer (NSCLC) sublines showed a collateral sensitivity to YTA0040. In transplantable murine tumor models, YTA0040 displayed a broad spectrum and high degree of anti-tumor activity when administered i.p. or p.o. (life span T/C = 135-234%). In P388 murine leukemia cells, YTA0040 caused dose- and time-dependent suppression of nucleic acid and protein synthesis, with protein synthesis being more potently and rapidly inhibited than nucleic acid synthesis. Flow-cytometric analysis revealed that YTA0040 blocked the entry of human NSCLC-derived A549 cells into S phase, leading to arrest in the G(1) phase of the cell cycle. Western blot analysis demonstrated that YTA0040 caused a dose-dependent decrease in the levels of expression of hyperphosphorylated pRb and cyclin A in A549 cells. The level of p53 protein expression was decreased by YTA0040 treatment. A higher concentration of YTA0040 down-regulated the levels of expression of CDK2, CDK4, cyclin D1 and cyclin E. These findings indicated that YTA0040 arrested human NSCLC cells in late G(1) phase of the cell cycle through inhibition of pRb phosphorylation. Inhibition of pRb phosphorylation by YTA0040 resulted from down-regulation of levels of expression of the CDKs and cyclins involved in the G(1)/S transition and not from induction of p53 and/or the CDK inhibitor p21., (Copyright 2000 Wiley-Liss, Inc.)

A tumor-suppressor gene, p16(INK4), which is deleted or mutated in tumors, regulates cell-cycle progression through a G(1)-S restriction point by inhibiting CDK4(CDK6)/cyclin-D-mediated phosphorylation of pRb. We have found that ectopic p16(INK4) expression increased cellular sensitivity of human non-small-cell-lung-cancer (NSCLC) A549 cells to a selective growth-inhibitory effect induced by the topoisomerase-I inhibitor 11, 7-ethyl-10-[4-(1-piperidino)-1-piperidino] carbonyloxy camptothecin (CPT-11) in vitro. In this study, we observed enhanced apoptosis characterized by DNA fragmentation in A549 cells transfected with p16(INK4) cDNA (A549/p16-1) and treated with CPT-11. This apoptosis was suppressed by the inhibitor of interleukin-1beta-converting enzyme (ICE/caspase-1) or ICE-like proteases, Z-Asp-CH2-DCB, as determined by DNA fragmentation and proteolytic cleavage of poly(ADP-ribose) polymerase, a natural substrate for CPP32/caspase-3. In A549/p16-1 cells, cytosolic peptidase activities that cleaved Z-DEVD-7-amino-4-trifluoromethylcoumarin increased during CPT-11-induced apoptosis and were suppressed by a highly specific caspase-3 and caspase-3-like inhibitor, Z-DEVD-fluoromethylketone. These findings indicate that p16(INK) is positively involved in the activation pathway of the caspase-3 induced by CPT-11. The increased delay in S-phase progression and subsequent induction of apoptosis were observed in CPT-11-treated A549/p16-1 cells on the basis of DNA histograms. Specific down-regulation of the cyclin-A protein level in A549/p16-1 cells was observed after CPT-11-treatment, whereas cyclin B, cdk2, and cdc2 protein levels were unaffected. These results suggest that ectopic p16(INK4) expression inappropriately decreases cyclin A and thereby terminates CPT-11-induced G(2)/M accumulation, which is followed by increased apoptosis in p16(INK4)-expressing A549 cells., (Copyright 2000 Wiley-Liss, Inc.)

UCN-01 (7-hydroxystaurosporine) inhibits the growth of various malignant cell lines in vitro and in vivo. In this study, a human small cell lung carcinoma subline resistant to UCN-01, SBC-3/UCN, was established and characterized. SBC-3/UCN cells showed 8-fold greater resistance to the UCN-01-induced growth-inhibitory effect than the parent cells, SBC-3. No UCN-01-induced G1 accumulation in SBC-3 cells was observed in SBC-3/UCN cells and decreased expression of phosphorylated RB protein was found in SBC-3 cells. Neither basal expression nor induction of p21(Cip1) by UCN-01 treatment was detected in the SBC-3/UCN cell line. An inhibitory effect of UCN-01 on CDK2 activity, which is mediated by p21(Cip1)/CDK2 complex formation upon UCN-01 treatment, was observed in SBC-3 but not in SBC-3/UCN cells. SBC-3/UCN showed higher CDK6 activity than SBC-3 cells. UCN-01 did not inhibit the CDK4 and CDK6 activities in both cells. We screened the cell cycle regulatory molecules associated with G(1)/S progression and found a remarked decrease in interferon regulatory factor 1 (IRF-1), which is known to cooperate with p53 in p21(Cip1) induction. Our results suggest that p21(Cip1) regulation via the IRF-1-associated pathway may represent a major determinant of UCN-01-induced growth inhibition in human lung cancer cells., (Copyright 2000 Wiley-Liss, Inc.)

Ganglioside GM2 is expressed on the surface of neuroblastoma and glioblastoma cells, and may also be detected on lung cancer cells. We reported previously that anti-ganglioside GM2 antibody exhibited strong in vitro anti-tumor activity against adriamycin-resistant cancer cells, which overexpressed ganglioside GM2. In the present study, we examined the in vivo anti-tumor effect of the chimeric anti-ganglioside GM2 antibody, KM966, against human lung and breast carcinoma cells, SBC-3 and MCF-7, and respective adriamycin-resistant clones, SBC-3/ADM and AdrR MCF-7 in BALB/c nu/nu mice. Ratios of tumor volume (T/C) between KM966-treated group and control group were 0.01 for SBC-3, 0.00 for SBC-3/ADM, 0.85 for MCF-7 and 0.34 for AdrR MCF-7 cells, respectively. Nude mice, which were pretreated with anti-asialo GM1 antibody to remove natural killer cells, were transplanted with 4 x 10(7) of SBC-3 and SBC-3/ADM subcutaneously. Seven days later, when tumors had grown to a diameter of over 8 mm, mice began to receive intravenous treatment of 120 microgram/mouse KM966 daily. Fourteen daily treatments induced regression to less than 4-mm diameter in 4/5 SBC-3 tumors and 5/5 of SBC-3/ADM tumors. All SBC-3/ADM tumors disappeared completely, suggesting that KM966 exerts a strong in vivo anti-tumor effect on ganglioside GM2-expressing cancer cells. In KM966-treated mice, the surface of the tumor cells stained positive with anti-human IgG. In addition, numerous leukocytes had infiltrated into the tumor mass. Antibody-dependent cell-mediated cytotoxicity (ADCC) of KM966 against tumor cells was examined in vitro by (51)Cr-release assay and revealed that KM966 induces ADCC activity against ganglioside GM2-expressing tumors. Our results suggest that immunotherapy using KM966 may be useful for the treatment of ganglioside GM2-expressing solid tumors., (Copyright 1999 Wiley-Liss, Inc.)

The induction and augmentation of tumor non-specific immunity and of tumor-specific immunity by intrapleural, intraperitoneal and subcutaneous injection of interleukin-2 (IL-2) gene-modified Lewis lung carcinoma (LLC) cells (LLC-IL2) was tested in C57BL/6 mice. Intrapleural injection of LLC cells induced lung tumors with a malignant effusion, intraperitoneal injection induced peritoneal tumors with ascites and subcutaneous injection induced subcutaneous tumors. Intrapleural injection of irradiated LLC-IL2 cured pre-existing lung LLC tumors and extended the survival of the mice but did not affect survival of mice with pre-existing peritoneal tumors nor did it affect the growth of s.c. tumors. Intraperitoneal injection of irradiated LLC-IL2 cured pre-existing LLC peritoneal tumors and extended the survival of the mice but did not affect survival of mice bearing lung tumors nor did it affect the growth of s.c. tumors. Subcutaneous injection of irradiated LLC-IL2 did not affect the growth of preexisting s.c. tumors and also did not improve survival of mice bearing the lung or peritoneal tumors. Injection with irradiated LLC-IL2 by all routes, i.e., intrapleural, intraperitoneal and s.c., protected against subsequent re-challenge with LLC. Eight days after the initial immunization (early stage of immunization), non-adherent mononuclear cells in the peritoneal cavity of the mice treated with intraperitoneal injection of irradiated LLC-IL2 displayed enhanced cytotoxicity against LLC, B16-F10 and P815 cells, while the cytotoxic activity of spleen cells in the same mice did not change. The efficiency of induction of tumor-specific immunity was the strongest after intraperitoneal immunization and weakest after s.c. immunization. In vitro analysis using the spleen cells of mice immunized with irradiated LLC-IL2 suggested that CD8+ T cells play a key role in tumor-specific immunity.

A novel antitumor antibiotic 7-N-[2-[[2-(gamma-L-glutamylamino)ethyl]dithio]ethyl] mitomycin C (KW-2149), an analogue of mitomycin C (MMC), is activated by thiol molecules, such as glutathione (GSH). To clarify the relationship between cellular GSH levels and the cytotoxicity of KW-2149, a murine fibroblast cell line (NIH/3T3) was transfected with human gamma-glutamylcysteine synthetase (gamma-GCS) cDNA, which codes a rate-limiting enzyme of GSH synthesis. Transfected cells (3T3/GCS) displayed increased gamma-GCS mRNA levels, gamma-GCS activity and GSH content, compared with NIH/3T3 cells. 3T3/GCS cells exhibited a 4.4-fold resistance to MMC, but not to KW-2149 (x 0.69), using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, suggesting that the increased cellular GSH levels did not affect the growth-inhibitory effect of KW-2149. KW-2149 exerted a greater growth-inhibitory effect than MMC on cisplatin- and doxorubicin-resistant cells with cross-resistance to MMC. KW-2149 exhibited a greater growth inhibitory effect than MMC not only on cells with GSH-mediated MMC resistance but also on cells with acquired resistance. We thus conclude that KW-2149 might be a clinically useful drug.

Ganglioside GM2 is one of the major cell-surface gangliosides expressed in human tumors. We earlier established a mouse/human IgG1 chimeric anti-GM2 antibody, KM966, which displayed anti-tumor activity in human tumor cells both in vitro and in vivo. In this study, we have screened for changes in ganglioside expressions in several drug-resistant human cancer cell lines to examine the modulation of drug resistance by immunotherapy with anti-ganglioside antibodies. Increased GM2 expression, detected by flow cytometry and thin-layer chromatography, was observed in the SBC-3/ADM and AdrR MCF7 adriamycin-resistant cell lines, in contrast with their parental lines. In other related gangliosides, ganglioside GD2 levels in AdrR MCF7 were higher than those in MCF7 cells. We confirmed increased N-acetylgalactosaminyltransferase mRNA in adriamycin-resistant cell lines, as compared with the parental cells, by Northern-blot analysis. Moreover, to investigate the possibility of exploiting the anti-tumor activity of KM966 in order to overcome resistance to adriamycin, we investigated the antibody-dependent cell-mediated cytotoxity of human peripheral mononuclear blood cells and the complement-dependent cytotoxity of human serum with KM966 against SBC-3, SBC-3/ADM, MCF7 and AdrR MCF7. Significantly higher killing via KM966 was observed in SBC-3/ADM and AdrR MCF7 cells as compared with the parental cells. This suggests that passive immunotherapy using KM966 against human adriamycin-resistant cancer may be useful for overcoming resistance to adriamycin.

Paclitaxel, an anti-mitotic anti-cancer agent, is active against solid tumors. The inhibition of depolymerization and promotion of microtubular assembly are essential for the anti-tumor activity of paclitaxel. Microtubule-associated proteins (MAPs) co-polymerize with tubulin and play some roles in microtubular dynamics. We examined the effect of paclitaxel on the interaction between tubulin and MAPs. Human lung-cancer cells, PC-14, were synchronized to G1/S border by the thymidine-double-block technique. After release from exposure to thymidine, the cells were treated briefly with 2 nM paclitaxel and the levels of alpha and beta tubulins and MAPs were examined after various times. Immunoblot analysis of paclitaxel-treated cells showed no changes in the overall expression of alpha and beta tubulins, microtubule-associated protein 2 (MAP2) or MAPs in comparison with controls. The samples were immunoprecipitated with anti-alpha- and anti-beta-tubulin antibodies and reblotted with an anti-MAP2 antibody, which showed that the amount of co-immuno-precipitated MAP2 in the synchronized cells, were increased by the brief paclitaxel treatment. These results suggest that paclitaxel treatment enhances the interaction between alpha and beta tubulins and MAP2. Since the phosphorylation state of MAP2 regulates the affinity of MAP2 for tubulins, and mitogen-activated protein (MAP) kinase is considered to be one of the kinases responsible for MAP2 phosphorylation, the effect of paclitaxel treatment on the MAP-kinase activity of synchronized PC-14 cells was examined. Two bands with molecular masses of 42 and 44 kDa were detected by an "intra-gel" MAP-kinase assay using myelin basic protein as the substrate. Paclitaxel treatment inhibited the MAP-kinase activity of PC-14 cells and inhibition was maximal at the G2/M phase of the cell cycle. Similar, concentration-dependent inhibition by paclitaxel of cellular MAP kinase of human synchronized small-cell lung carcinoma, H69, was observed. No inhibition of the MAP kinase of the paclitaxel-resistant sub-line H69/Txl by paclitaxel was observed, suggesting that some change of the MAP-kinase cascade had occurred in these cells. No direct inhibition of MAP-kinase activity by paclitaxel was observed in the cell-free assay (in vitro), suggesting that paclitaxel did not inhibit MAP kinase directly. Since it has been speculated that p34cdc2 kinase is also a kinase that phosphorylates MAP2, the effect of paclitaxel treatment on the p34cdc2-kinase activity of synchronized PC-14 and PC-9 cells was examined. Paclitaxel inhibited p34cdc2-kinase activation at the G2/M phase. These results suggest that paclitaxel inhibited MAP kinase and p34cdc2 kinase in vivo indirectly. These actions of paclitaxel may be responsible for the increased affinity between MAP2 and tubulins that it induces.

A subline of rat C6 glioma cells, C6-10A cells, in which epinephrine can induce nerve growth factor (NGF) synthesis/secretion, was isolated. C6-10A cells have retained their sensitivity to epinephrine for more than 2 years in a medium containing 0.5% fetal calf serum (FCS) but easily lose it in 10% FCS. C6-10A cells are S-100- and glial fibrillary acidic protein-positive, and the doubling time is about 60 h in the medium containing 0.5% FCS and about 20 h in 10% FCS. Epinephrine induced NGF synthesis/secretion prominently in serum-free cultures of C6-10A cells and in cultures with a high cell density, but not in serum-containing cultures. The induction did not occur with parent C6 cells under the appropriate conditions in C6-10A cells. NGF secretion was induced by catecholaminergic compounds in the following order isoproterenol > epinephrine = norepinephrine >> dopamine. The induction caused by epinephrine was blocked by propranolol (beta-blocker) but not by phentolamine (alpha-blocker). Various compounds that activate the adenylate cyclase system also induced NGF synthesis/secretion. These results indicate that C6-10A cells are astrocytes that are highly responsive to beta-adrenergic receptor agonists, which stimulate NGF synthesis/secretion via receptors coupled with adenylate cyclase machinery. These cells may be a useful aid in studying the mechanism of NGF synthesis/secretion.