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1.
Yamamoto N Suyama H Nakazato H Yamamoto N Koga Y 《Cancer immunology, immunotherapy : CII》2008,57(7):1007-1016
Serum vitamin D binding protein (Gc protein) is the precursor for the principal macrophage-activating factor (MAF). The MAF
precursor activity of serum Gc protein of colorectal cancer patients was lost or reduced because Gc protein is deglycosylated
by serum α-N-acetylgalactosaminidase (Nagalase) secreted from cancerous cells. Deglycosylated Gc protein cannot be converted
to MAF, leading to immunosuppression. Stepwise treatment of purified Gc protein with immobilized β-galactosidase and sialidase
generated the most potent macrophage-activating factor (GcMAF) ever discovered, but it produces no side effect in humans.
Macrophages treated with GcMAF (100 pg/ml) develop an enormous variation of receptors and are highly tumoricidal to a variety
of cancers indiscriminately. Administration of 100 nanogram (ng)/human maximally activates systemic macrophages that can kill
cancerous cells. Since the half-life of the activated macrophages is approximately 6 days, 100 ng GcMAF was administered weekly
to eight nonanemic colorectal cancer patients who had previously received tumor-resection but still carried significant amounts
of metastatic tumor cells. As GcMAF therapy progressed, the MAF precursor activities of all patients increased and conversely
their serum Nagalase activities decreased. Since serum Nagalase is proportional to tumor burden, serum Nagalase activity was
used as a prognostic index for time course analysis of GcMAF therapy. After 32–50 weekly administrations of 100 ng GcMAF,
all colorectal cancer patients exhibited healthy control levels of the serum Nagalase activity, indicating eradication of
metastatic tumor cells. During 7 years after the completion of GcMAF therapy, their serum Nagalase activity did not increase,
indicating no recurrence of cancer, which was also supported by the annual CT scans of these patients. 相似文献
2.
Jong Young Choi Su Hee Lee Hyon Bin Na Kwangjin An Taeghwan Hyeon Tae Seok Seo 《Bioprocess and biosystems engineering》2010,33(1):21-30
In this study, we present in vitro cytotoxicity of iron oxide (Fe3O4) and manganese oxide (MnO) using live/dead cell assay, lactate dehydrogenase assay, and reactive oxygen species detection
with variation of the concentration of nanoparticles (5–500 μg/ml), incubation time (18–96 h), and different human cell lines
(lung adenocarcinoma, breast cancer cells, and glioblastoma cells). The surface of nanoparticles is modified with polyethyleneglycol-derivatized
phospholipid to enhance the biocompatibility, water-solubility, and stability under an aqueous media. While the cytotoxic
effect was negligible for 18 h incubation even at highest concentration of 500 μg/ml, MnO nanoparticle represented higher
level of toxicity than those of Fe3O4 and the commercial medical contrast reagent, Feridex after 2 and 4 day incubation time. However, the cytotoxicity of Fe3O4 is equivalent or better than Feridex based on the live/dead cell viability assay. The engineered MnO and Fe3O4 exhibited excellent stability compared with Feridex for a prolonged incubation time. 相似文献
3.
Suhrid Banskota Jaya Gautam Sushil C. Regmi Pallavi Gurung Myo-Hyeon Park Seung Joo Kim Tae-gyu Nam Byeong-Seon Jeong Jung-Ae Kim 《PloS one》2016,11(1)
5-Hydroxytryptamine (5-HT) induces proliferation of cancer cells and vascular cells. In addition to 5-HT production by several cancer cells including gastrointestinal and breast cancer, a significant level of 5-HT is released from activated platelets in the thrombotic environment of tumors, suggesting that inhibition of 5-HT signaling may constitute a new target for antiangiogenic anticancer drug discovery. In the current study we clearly demonstrate that 5-HT-induced angiogenesis was mediated through the 5-HT1 receptor-linked Gβγ/Src/PI3K pathway, but not through the MAPK/ERK/p38 pathway. In addition, 5-HT induced production of NADPH oxidase (NOX)-derived reactive oxygen species (ROS). In an effort to develop new molecularly targeted anticancer agents against 5-HT action in tumor growth, we demonstrate that BJ-1108, a derivative of 6-amino-2,4,5-trimethylpyridin-3-ol, significantly inhibited 5-HT-induced angiogenesis. In addition, BJ-1108 induced a significant reduction in the size and weight of excised tumors in breast cancer cell-inoculated CAM assay, showing proportionate suppression of tumor growth along with inhibition of angiogenesis. In human umbilical vein endothelial cells (HUVECs), BJ-1108 significantly suppressed 5-HT-induced ROS generation and phosphorylation of PI3K/Akt but not of Src. Unlike NOX inhibitors, BJ-1108, which showed better antioxidant activity than vitamin C, barely suppressed superoxide anion induced by mevalonate or geranylgeranyl pyrophosphate which directly activates NOX without help from other signaling molecules in HUVECs, implying that the anti-angiogenic action of BJ-1108 was not mediated through direct action on NOX activation, or free radical scavenging activity. In conclusion, BJ-1108 inhibited 5-HT-induced angiogenesis through PI3K/NOX signaling but not through Src, ERK, or p38. 相似文献
4.
Maria-Eleni Androutsou Mahmoud Saifeddine Morley D. Hollenberg John Matsoukas George Agelis 《Amino acids》2010,38(4):985-990
In the present study, we report the synthesis and biological evaluation of a series of new non-peptide PAR1 mimetic receptor antagonists, based on conformational analysis of the S42FLLR46 tethered ligand (TL) sequence of PAR1. These compounds incorporate the key pharmacophore groups in the TL sequence, guanidyl, amino and phenyl, which are essential
for triggering receptor activity. Compounds 5 and 15 (50–100 μM) inhibited both TFLLR-amide (10 μM) and thrombin-mediated (0.5 and 1 U/ml; 5 and 10 μM) calcium signaling in a
cultured human HEK cell assay. 相似文献
5.
Hsiao YC Hsieh YS Kuo WH Chiou HL Yang SF Chiang WL Chu SC 《Journal of biomedical science》2007,14(1):107-119
Summary Natural products, including flavonoids, are suggested to be involved in the protective effects of fruits and vegetables against
cancer. However, studies concerning the effect of flavonoids frequently lacked data regarding to flavanones. In this study,
we investigated the inhibitory effect of flavanone compounds, including flavanone, 2′-OH flavanone, 4′-OH flavanone, 6-OH
flavanone, naringin and naringenin, on cell growth of various cancer cells. We determined that flavanone and 2′-OH flavanone
inhibited cell growth of A549, LLC, AGS, SK-Hepl and HA22T cancer cells, while other flavanones showed little or no inhibition.
We evaluated growth-inhibitory activity of flavanone and 2′-OH flavanone against highly proliferative human lung cancer cells
(A549) via anchorage-independent and -dependent colony formation assay, and further showed that treatment of flavanone resulted
in a G1 cell cycle arrest with reduction of cyclin D, E and cyclin-dependent kinase (CDK) 2, while treatment of 2′-OH flavanone
led to a G2/M phase accumulation with reduction of cyclin B, D and Cdc2. Moreover, we demonstrated the improvement effect
of flavanone and 2′-OH flavanone with anti-cancer drug, doxorubicin, on A549 cells. Finally, flavanone and 2′-OH flavanone
were evidenced by its inhibition on the growth of A549 and Lewis lung carcinoma cells in vivo.
Yung-Chin Hsiao and Yih-Shou Hsieh are equally contributed to this work. 相似文献
6.
Kuniyo Inouye 《Cancer immunology, immunotherapy : CII》1997,45(3-4):159-161
A bispecific F(ab′)2 fragment recognizing both human thyroid-stimulating hormone (TSH) and alkaline phosphatase (AP) was prepared by disulfide
bond exchange between F(ab′)2 fragments of IgG1 mAb against TSH and AP. AP was polymerized by glutaraldehyde, and a sandwich enzyme-linked immunosorbent
assay for TSH was developed by using the AP polymers and the bispecific F(ab′)2 fragment. In this assay, the preparation of covalently linked AP-mAb conjugates was not needed, and the interaction of mAb
with non-specific proteins was greatly reduced. The sensitivity for TSH increased in proportion to the degree of AP polymerization,
and the lower detection limit obtained with the AP trimer was 0.5 μU/ml. The use of the bispecific F(ab′)2 fragment allows us to use monomers and polymers of AP and thereby regulates the sensitivity of the assay.
Accepted: 14 October 1997 相似文献
7.
Sophie Bombard Marzia Bruna Gariboldi Elena Monti Elisabetta Gabano Luca Gaviglio Mauro Ravera Domenico Osella 《Journal of biological inorganic chemistry》2010,15(6):841-850
Enantiomeric complexes of formula [PtCl2L2] [L2 is (R)-(+)-BINAP and (S)-(−)-BINAP, where BINAP is 2,2′-bis(diphenylphosphane)-1,1′-binaphthyl, and (R)-(+)-DABN and (S)-(−)-DABN, where DABN is 1,1′-binaphthyl-2,2′-diamine], were tested for their cytotoxic activity against three cancer cell
lines and for their ability to bind to the human telomeric sequence folded in the G-quadruplex structure. Similar experiments
were carried out on prototypal complexes cisplatin and cis-[PtCl2(PPh3)2] for comparison. Platinum complexes containing phosphanes proved less cytotoxic to cancer cell lines and less likely to interact
with the nucleobases of the G-quadruplex than those containing amines; in both cases the S-(−) isomer was more active than the R-(+) counterpart. More specifically, whereas all the platinum complexes were able to platinate the G-quadruplex structure
from the human telomeric repeat, the extent and sites of platination depended on the nature of the ligands. Complexes containing
(bulky) phosphanes interacted only with the adenines of the loops, whereas those containing the less sterically demanding
amines interacted with adenines and some guanines of the G-quartet. 相似文献
8.
Pavla Heringova Jana Kasparkova Viktor Brabec 《Journal of biological inorganic chemistry》2009,14(6):959-968
We studied the effect of antitumor cisplatin and inefficient transplatin on the structure and stability of G quadruplexes
formed by the model human telomere sequence 5′-GGG(TTAGGG)3-3′ using circular dichroism, UV-monitored thermal denaturation, and gel electrophoresis. In addition, to investigate whether
there is a connection between the ability of cisplatin or transplatin to affect telomerase activity and stability of G quadruplexes,
we also used a modified telomere repeat amplification protocol assay that uses an oligonucleotide substrate for telomerase
elongation susceptible to forming a G quadruplex. The results indicate that cisplatin is more efficient than transplatin in
disturbing the quadruplex structure, thereby precluding telomeric sequences from forming quadruplexes. On the other hand,
the results of this work also demonstrate that in absence of free platinum complex, DNA adducts of antitumor cisplatin inhibit
telomerase catalysis, so the mechanism underlying this inhibition does not involve formation of the G quadruplexes which are
not elongated by telomerase. 相似文献
9.
Influence of stereochemistry of the 3′-terminal phosphorothioate (PS)-modified primers was studied in a single base extension
(SBE) assay to evaluate any improvements in specificity. SBE reactions were catalyzed by members of the high fidelity Pfu family of DNA polymerases with (exo+) or without (exo−) 3′ → 5′ exonucleolytic activity. The diastereomerically pure PS-labeled
primers used in these studies were obtained either by the stereospecific chemical synthesis invented in our laboratory or
by the more conventional ion-exchange chromatographic method for separation of a mixture of diastereomers (RP and SP). When the SBE reaction was performed in the presence of mispaired 2′-deoxyribonucleoside triphosphates (dNTPs), the “racemic”
3′-phosphorothioate primer mixture resulted in a lower level of 3′ → 5′ exonuclease-mediated cleavage products in comparison
to the SBE reactions carried out with the corresponding unmodified primer. When the diastereomerically pure RP 3′-phosphorothioate primer was examined, the results were largely the same as for the racemic 3′-phosphorothioate primer
mixture. In contrast, a 3′-PS primer of SP configuration displayed significantly improved performance in the SBE reaction. This included the lack of 3′ → 5′ proofreading
products, less mispriming, and improved yield of incorporation of the correct nucleotide. 相似文献
10.
Diamantis Konstantinidis Konstantinos Paletas George Koliakos Martha Kaloyianni 《Cell stress & chaperones》2009,14(6):591-601
Leptin, a 16-kDa cytokine produced mainly by the adipose tissue, is known to increase energy expenditure while at the same
time lowering food intake by acting directly on the hypothalamus. ObRb, the leptin receptor mostly involved in intracellular
signaling, is expressed in a wide range of tissues, thus allowing leptin to affect a much broader diversity of biological
processes. High concentrations of leptin are encountered in patients with hyperleptinemia, a condition which very often accompanies
obesity and which is a direct result of leptin resistance. In the present study, moderate and high concentrations of leptin
(16 and 160 ng/ml) were mostly utilized in order to investigate the role of this cytokine in oxidative stress levels in human
monocytes. Leptin was found to increase oxidative species production as measured with 2′,7′-dichlorodihydrofluorescein diacetate
(general marker of oxidative species, but not O2−.) and dihydroethidium (marker of O2−.). Surprisingly, it also augmented superoxide dismutase activity. Inhibition of the Na+–H+ exchanger isoform 1 (NHE1) also inhibited leptin-induced superoxide anion production but at the same time amplified leptin-induced
production of other oxidative species. Signaling proteins such as phosphoinositide 3 kinase and conventional isoforms of protein
kinase C (α-, βi-, βii-), as well as NADPH oxidase, also participated in leptin signaling. Finally, leptin was found to increase glutathionylation
levels of NHE1-bound heat shock protein 70 kDa (Hsp70) but not Hsp70 binding to NHE1. 相似文献
11.
Two zinc(II) terpyridine complexes Zn(atpy)2(PF6)2 (1) (atpy = 4′-p-N9′-adeninylmethylphenyl-2,2′:6,2′′-terpyridine) and Zn(ttpy)2(PF6)2 (2) (ttpy = 4′-p-tolyl-2,2′:6,2′′-terpyridine) have been synthesized and characterized by elemental analysis, 1H NMR and electrospray mass spectroscopy. The structure of complex 2 was also determined by X-ray crystallography, which revealed a ZnN6 coordination in an octahedral geometry with two terpyridine acting as equatorial ligands. The circular dichroism data showed
that complex 1 exhibited an ICD signal at around 300 nm and induced more evident disturbances on DNA base stacking than complex 2, reflecting the impact of the adenine moiety on DNA binding modes. Complex 1 exhibited higher cleavage activity to supercoiled pUC 19 DNA than complex 2 under aerobic conditions, suggesting a promotional effect of adenine moiety in DNA nuclease ability. Interestingly, both
complexes demonstrated potent in vitro cytotoxicity against a series human tumor cell lines such as human cervix carcinoma
cell line (HeLa), human liver carcinoma cell line (HepG2), human galactophore carcinoma cell line (MCF-7) and human prostate
carcinoma cell line (pc-3). The cytotoxicity is averagely 10 times more active than the anticancer drug cisplatin.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
12.
Scorei R Ciubar R Ciofrangeanu CM Mitran V Cimpean A Iordachescu D 《Biological trace element research》2008,122(3):197-205
Recent studies suggested that boron has a chemo-preventive role in prostate cancer. In the present report, we investigated
the effects of calcium fructoborate (CF) and boric acid (BA) on activation of the apoptotic pathway in MDA-MB-231 human breast
cancer cells. Exposure to BA and CF inhibited the proliferation of breast cancer cells in a dose-dependent manner. Treatment
with CF but not BA resulted in a decrease in p53 and bcl-2 protein levels. Furthermore, after the treatment with CF, augmentation
of pro-caspase-3 protein expression, cytosolic cytochrome c level, and caspase-3 activity were observed, indicating apoptotic cell death induction. This was also demonstrated by terminal
deoxynucleotidyl transferase-mediated 2′-deoxyuridine 5′-triphosphate nick-end-labeling assay. In conclusion, our data provide
arguments to the fact that both BA and CF inhibited the growth of breast cancer cells, while only CF induced apoptosis. Additional
studies will be needed to identify the underlying mechanism responsible for the observed cellular responses to these compounds
and to determine if BA and CF may be further evaluated as chemotherapeutic agents for human cancer. 相似文献
13.
Irene van den Broek Rolf W. Sparidans Jan H.M. Schellens Jos H. Beijnen 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》2010,878(5-6):590-602
An assay to quantify several possible breast cancer peptide biomarkers in human serum has been developed and validated, using liquid chromatography coupled to tandem mass spectrometry (LC–MS/MS). The peptides include bradykinin, Hyp3-bradykinin, des-Arg9-bradykinin and fragments of fibrinogen α-chain (Fib-α[605–629]), inter-α-trypsin inhibitor heavy chain 4 (ITIH4[666–687]) and complement component 4a (C4a[1337–1350]). Ile13-ITIH4[666–687], d20-C4a[1337–1350] and Sar-D-Phe8-des-Arg9-bradykinin were used as internal standards. Bovine plasma, with 2 mM captopril and 2 mM d-l-mercaptoethanol-3-guanidino-ethylthiopropanoic acid (MEGETPA) to prevent rapid degradation of the bradykinins, was used as analyte-free matrix. Recoveries for solid-phase extraction (SPE) on mixed-mode weak cation exchange sorbents were between 62 and 90%. Multiple reaction monitoring (MRM) on a triple quadrupole mass spectrometer equipped with a heated electrospray source (H-ESI), operating in the positive ion-mode, was used for detection. The assay was fully validated and stabilities of the peptides were extensively explored. Bradykinin (10–500 ng/ml), Hyp3-bradykinin (4–200 ng/ml), des-Arg9-bradykinin (2–100 ng/ml), Fib-α[605–629] (120–3000 ng/ml), ITIH4[666–687] (0.4–10 ng/ml) and C4a[1337–1350] (1–25 ng/ml) were simultaneously quantified with deviations from the nominal concentrations below 22% and intra- and inter-assay precisions below 15 and 20%, respectively, for all peptides at all concentrations. The method has been successfully applied to several serum samples from breast cancer patients and matched controls. 相似文献
14.
Ai-Lian Zhang Tian-Yuan Zhang Jin-Xian Luo Ce-Yi Fu Zhi Qu Guo-Hui Yi Dong-Xiao Su Fa-Zhi Tu Ying-Wen Pan 《Molecular biology reports》2009,36(8):2265-2270
A high-density cell culture method was successfully established in P. pastoris with the alcohol oxidase I (AOXI) promoter in order to produce large quantities of recombinant human angiostatin (AS) which
has been reported to have antiangiogenic activity. A preliminary study on fermentation conditions in shaking flasks indicated
that adequacy of biomass is beneficial to obtain more products. The fermentation was carried out in a 10 l bioreactor with
5 l modified growth medium recommended by Invitrogen at 30°C. The cells were first grown in glycerol-PTM4 trace salts for
24 h. When the cell density reached A600 = 125, methanol-PTM4 trace salts was added to induce the expression of AS. During the fermentation, dissolved oxygen level
was maintained at 20–30%, pH was controlled at 5 by the addition of 7 M NH4OH and the biomass was maintained at about A600 = 200. After 60 h of induction, the secreted AS was 153 mg/l. The recombinant AS inhibited the angiogenesis on CAM and suppressed
the growth of B16 melanoma in C57BL/6J mice (P < 0.01). 相似文献
15.
Nagai Y Tasaki H Takatsu H Nihei S Yamashita K Toyokawa T Nakashima Y 《Biochemical and biophysical research communications》2001,281(3):726-731
Homocysteine has been reported to inhibit endothelial cell proliferation, which is closely related to angiogenesis. However, the relationship between homocysteine and angiogenesis is unknown. To clarify whether homocysteine would inhibit angiogenesis in vitro and in vivo, we examined the effect of homocysteine on tube formation by bovine aortic endothelial cells (BAECs) and by human microvessel endothelial cell-1 (HMEC-1) in vitro, and on angiogenesis in vivo using the chorioallantoic membrane (CAM) assay, as well as on BAEC proliferation and migration. Homocysteine, but not cysteine, inhibited BAEC proliferation, migration, and tube formation in a dose-dependent manner at concentrations from 0 to 10 mM. Homocysteine also inhibited tube formation by HMEC-1s. In these assay, 50% inhibition was induced by about 1 mM homocysteine. In the in vivo CAM assay, 0, 10, 100, 500, and 1000 microgram homocysteine induced an avascular zone by 0, 0, 16.7, 53.3 and 76.5%, respectively, also showing a dose-dependent effect. It was suggested that homocysteine inhibited angiogenesis by preventing proliferation and migration of endothelial cells. 相似文献
16.
The increasing applications of silicon dioxide (SiO2) nanomaterials have been widely concerned over their biological effects and potential hazard to human health. In this study,
we explored the effects of SiO2 nanoparticles (15, 30, and 100 nm) and their micro-sized counterpart on cultured human epidermal Keratinocyte (HaCaT) cells.
Cell viability, cell morphology, reactive oxygen species (ROS), DNA damage (8-OHdG, γH2AX and comet assay) and apoptosis were
assessed under control and SiO2 nanoparticles exposed conditions. As observed in the Cell Counting Kit-8 (CCK-8) assay, exposure to 15, 30 or 100 nm SiO2 nanoparticles at dosage levels between 0 and 100 μg/ml decreased cell viability in a concentration- and size dependent manner
and the IC50 of 24 hour exposure was 19.4 ± 1.3, 27.7 ± 1.5 and 35.9 ± 1.6 μg/ml for 15, 30 and 100 nm SiO2 nanoparticles, respectively. Morphological examination revealed cell shrinkage and cell wall missing after SiO2 nanoparticle exposure. Increase in intracellular ROS level and DNA damage as well as apoptosis were also observed in SiO2 nanoparticle-exposed HaCaT cells. Exposure to SiO2 nanoparticles results in a concentration- and size-dependent cytotoxicity and DNA damage in cultural HaCaT cells which is
closely correlated to increased oxidative stress. 相似文献
17.
The functional Mn content of intact photosystem II membrane fragments was measured as 4.06 ± 0.13 Mn/reaction center when
determined using a simple, sensitive colorimetric assay that will also work with thylakoids and core complexes. This procedure
requires minimal sample material, does not need expensive assay equipment, requires four simple steps, and only takes 20–30 min
to perform. These include (a) removal of the adventitious Mn ions by CaCl2 treatment of the membranes, (b) extraction of the Mn from the O2-evolving complex with hydrochloric acid, (c) purification of the extract by centrifugation followed by filtration of the
supernatant through an Acrodisc syringe filter (0.2 μm nylon membrane), and (d) colorimetric determination of Mn in the extract
using the reaction of the chromogenic agent, 3,3′,5,5′-tetramethylbenzidine, with previously oxidized Mn(II) cations carried
out at high pH. The colorimetric assay itself has been used previously by Serrat (Mikrochim Acta 129:77–80, 1998) for assaying
Mn concentrations in sea water and drinking water. 相似文献
18.
Pesticidal activity of Bacillus thuringiensisδ-endotoxins, Cry1Aa, Cry1Ab, Cry1Ac, and Cry2A, was determined by using the force-feeding bioassay method to 4th instar larvae of Helicoverpa zea. H. zea was susceptible to Bt toxins in the order Cry1Ac > Cry1Ab > Cry1Aa > Cry2A with 63.60, 89.04, 159.65, and 375.78 ng/larvae
respectively. The abilities of selected Bacillus thuringiensis toxins to inhibit short circuit current (ISC) in midgut epithelia of H. zea were also investigated by voltage clamp assay. The voltage-clamp studies were conducted on isolated midguts, measuring the
inhibition of short circuit current (ISC) by activated toxin. A Cry1Aa toxin dilution of 33.3 and 500 ng/ml resulted in inhibition of ISC of −2.29 μA/min (lag time 15 min) and −4.48 μA/min (lag time, 2 min) respectively. The Cry1Ab dilution of 25 ng/ml inhibited
ISC to −1.39 μA/min, a lag time of 14 min, and 333.3 ng/ml dilution resulted in decay of ISC−2.49 μA/min, lag time 1 min respectively. The Cry1Ac lower dilution 16.7 ng/ml inhibited ISC to −1.39 μA/min, lag time 4 min, and a high dilution 333.3 ng/ml decay ISC to −2.44 μA/min, lag time 1 min. The inhibition of ISC (−1.10 μA/min, lag time 25) at lower dilution (33.3 ng/ml) and high dilution (500 ng/ml), decay (−2.38 μA/min, lag time 5
min), showed a correlation between toxin concentration and inhibitory response with Cry2A toxin. The lag time decreased with
increasing concentration of toxin applied, which is additional evidence of dose response besides direct correlation of toxicity
assays and ISC.
Received: 7 April 2000 / Accepted: 2 May 2000 相似文献
19.
Terry L. Riss Kenneth P. Karey B. Daniel Burleigh David Farker David A. Sirbasku 《In vitro cellular & developmental biology. Plant》1988,24(11):1099-1106
Summary A serum-free clonal density growth assay was developed for the quantification of the biological activity of human recombinant
insulin-like growth factor I (IGF-I). The assay measures IGF-I stimulated growth of Balb/c 3T3 cells cultured over 4 d on
poly-d-lysine-coated plastic surfaces in a serum-free medium formulation composed of a 1∶1 (vol/vol) mixture of Ham's F12 and Dulbecco's
modified Eagle's media, supplemented with 3.0 ng/ml bovine basic fibroblast growth factor (bFGF), 10 μg/ml human transferrin,
100 μg/ml ovalbumin, and 1.0 μM dexamethanose. Low-temperature trypsinization of serum-supplemented stock cultures combined with the use of poly-d-lysine-coated plates made it unnecessary to use serum or fibronectin to promote cell attachment and survival. Serum-free
growth conditions were optimized with respect to the concentrations of the supplements. Addition of IGF-I resulted in 3.5-fold
more cells than control cultures without IGF-I after 4 d. Deletion of bFGF resulted in no IGF-I stimulation of growth. The
concentrations of various preparations of IGF-I required to achieve one-half maximal stimulation of cell number (ED50), ranged between 1.25 and 4.7 ng/ml. In parallel assays, IGF-I was 6.6 times more potent than human recombinant insulin-like
growth factor II and 32 times more potent than insulin. When cells were seeded into medium containing IGF-I, transferrin,
ovalbumin, and dexamethasone but no bFGF, growth was minimal. Dose-response addition of bFGF showed an ED50, of 0.9 ng/ml. The methods reported are useful to monitor the biological potency of recombinant and natural-source growth
factors as well as providing a new means of studying the multiple growth factor requirements of Balb/c 3T3 cells in cultures.
This work was supported by a contract from IMCERA Bioproducts, Inc. 相似文献
20.
O. M. Rozhmanova N. Kh. Pogorelaya E. V. Dolgaya L. N. Stelmakh I. S. Magura I. O. Mikhailopulo 《Neurophysiology》1998,30(4-5):250-252
We investigated the actions of human recombinant α2-interferon and a secondary messenger of its action, 2′-5′-oligoadenylate, on tetrodotoxin (TTX)-sensitive sodium transport
using human (IMR-32) and murine (NIE-115) neuroblastoma cells. In biochemical experiments using22Na, human interferon was shown to increase entry of22Na into IMR-32 neuroblastoma cells through the channels activated by veratrine and scorpion toxin. This increase was clearly
dose-dependent. Cell treatment with TTX completely inhibited this sodium transport. On the contrary, 2′-5′-oligoadenylate
depressed entry of22Na into neuroblastoma cells. The activation effect was not observed under the action of human α2-interferon on TTX-sensitive sodium flows to the murine neuroblastoma cells, which demonstrated the species-related specificity
of this agent. 相似文献