Roles of cytosolic phospholipase A(2) and platelet-activating factor receptor in the Ca-induced biosynthesis of PAF

Casein-elicited peritoneal exudate cells (PEC), mainly consisted of neutrophils, were collected from platelet-activating factor receptor-knock-out (PAFR-KO), cytosolic phospholipase A(2) knock-out (cPLA(2)-KO), and wild-type (WT) mice. After stimulation of PEC with calcium ionophore A 23187, PAF levels were measured by radio-ligand binding assay using receptor-rich membrane fraction prepared from the PAF receptor transgenic mice. We found that the level of PAF production by PEC was not different between WT and PAFR-KO mice. On the other hand, cPLA(2)-KO mice were deficient in the PAF production. These results provide the direct evidence while cPLA(2) is essential in the production of PAF, PAF receptor deficiency has little effect on the PAF production.     

The modelling of the growth of Ehrlich carcinoma and teratoma T-36 with ascitic fluid and tumor-cell dialysate]

The study of the effect of ascitic fluid and dialysate of Ehrlich ascites tumor cells (M.m. less than 15 kDa) on the growth of Ehrlich carcinoma and teratoma T-36 has shown that both the ascitic fluid and dialysate can protect tumor cells in vivo. The number of animals with tumors increased from 0% in control animals to 60 and 20%, respectively, in experimental ones after transplantation i.m. of 20 x 10(3) Ehrlich tumor cells into mice. Compared to control, ascitic fluid and dialysate of Ehrlich ascites tumor cells increased the rate of tumor growth to 195 and 153%, respectively. It is suggested that this test-system simulates the effect of tumor humoral factors in vivo.     

Increases in mRNA levels of glucose transporters types 1 and 3 in Ehrlich ascites tumor cells during tumor development

A common feature of many tumors is an increase in glucose catabolism during tumor growth. We studied the mechanism of this phenomenon by using Ehrlich ascites tumor bearing mice as the animal model. We found that Ehrlich ascites tumor cells possess only glucose transporter 1 (GLUT1) and GLUT3 but no GLUT2, GLUT4, or GLUT5. The mRNA levels of GLUT1 and GLUT3 increased progressively in the tumour during development; however, there were no changes observable in mRNA levels of glucose transporters of all types in brain, liver, and heart of the host mice. These findings suggest that Ehrlich ascites tumor augments its glucose transport mechanism relative to other tissues in response to its unique growth needs. J. Cell. Biochem. 67:131–135, 1997. © 1997 Wiley-Liss, Inc.     

Tumor-Specific Transplantation Resistance in Mice after Treatment of Initial Tumors with Streptococcus thermophilus

Antitumor activity observed by treatment with Streptococcus thermophilus was further investigated. The mice cured from fibrosarcoma by treatment with heat-killed preparation of S. thermophilus, when challenged with fibrosarcoma failed to take up the tumor. However, these cured mice when challenged with sarcoma-180 or Ehrlich ascites carcinoma, did not show significant changes in tumor take and/or survival compared to their respective controls. Similarly, mice cured from sarcoma-180 were challenged with fibrosarcoma, sarcoma-180 or Ehrlich ascites carcinoma. Though there was no change in the mean survival time (MST) of the dying mice regarding sarcoma-180 or Ehrlich ascites carcinoma, there was 50 and 30% increase in the number of mice that showed total regression respectively over controls. However, there was no difference in the growth rate of fibrosarcoma. Similar observations were made with mice cured from Ehrlich ascites carcinoma, challenged with these tumors. These findings thus suggest that the antitumor response was tumor-specific and that tumor-associated antigens may have a role in imparting this specificity. Bacterial treatment non-specifically augmented this primary response.     

Angiogenesis and inflammation in skeletal muscle in response to ascites tumor in mice

This study addresses the interaction between Ehrlich ascites tumor and skeletal abdominal muscle, presenting quantitative analysis of ascites-induced angiogenesis and inflammation in this tissue of mice bearing-tumor. Time-dependent changes in the muscle (cellular activity, angiogenesis, inflammation and cytokines production) were assessed by morphometric, functional, and biochemical parameters at days 1, 4 and 8 after i.p. inoculation of Ehrlich tumor cells (2.5 x 10(7)). The number of cells stained with AgNOR technique (argyrophilic nucleolar organizer region) in the muscle, together with MTS assay used as markers of cellular activity increased progressively in parallel with the out flow rate of sodium fluorescein (blood flow index), hemoglobin content (vascular index) and VEGF production. Likewise, the inflammatory process in the muscle, as assessed by myeloperoxidase (MPO) and n-acethylglucosaminidase (NAG) activities and the levels of the chemokines, keratinocyte-derived chemokine (CXC1-3/KC) and macrophage-chemoattractant protein (CCL2/MCP-1) increased with tumor development. The combination of techniques used to describe angiogenesis and inflammation in a muscle model system has proved to be suited for quantitative measurements of microvascular changes and cellular infiltration occurring in the abdominal muscle wall of ascites-bearing mice. This study holds potential for investigating events and mechanisms associated with skeletal muscle response to neoplasic stimulus.     

Effect of the diabetic environment on the lipid metabolism of Ehrlich ascites cells

One strain of Ehrlich ascites cells lacking of insulin receptors, was grown into control and diabetic mice and cells harvested from diabetic mice reimplanted into control mice. The fatty acid composition of neutral and polar lipids was analyzed and several parameters calculated. Results showed that it is possible to produce similar changes in the lipid fatty acid unsaturation of Ehrlich cells to those observed in the liver of the diabetic bearing mice. These changes may be reverted by growing these cells into control mice. The diabetic environment also promoted a relative increase in the radioactivity from incorporated in vitro into neutral lipids of Ehrlich cells. This metabolic adjustment, probably due to an induction of the enzyme diglyceride acyltransferase, was completely reverted by transplanting these cells in control mice. The metabolic adaptation of Ehrlich ascites cells to the diabetic environment did not modify their biological behaviour as pointed out by their mean generation time. The evidence presented here, showing relatively normal growth of Ehrlich cells in association with changes in the lipid fatty acid pattern and in lipid metabolism, indicates the adaptation of these cells, lacking of insulin receptors, to the environment provided by the diabetic mice.     

Intracranial microenvironment reveals independent opposing functions of host alphaVbeta3 expression on glioma growth and angiogenesis

alphaVbeta3 integrins are overexpressed in the host-derived vasculature of glioblastoma multiform (GBM) and are believed to contribute to angiogenesis and tumor growth. To directly address the role of host alphaVbeta3 expression in GBM growth and behavior, we intracranially implanted integrin beta3-expressing GBM cells into beta3 wild type (WT) or beta3 knock out (KO) mice and monitored angiogenesis and growth. GBM in beta3 WT animals had a vessel density greater than that in beta3 KO animals, consistent with a pro-angiogenic, pro-tumorigenic view of host integrin function. GBM in beta3 WT animals, however, were no larger than those in beta3 KO animals, because GBM in beta3WT animals were infiltrated with a higher number of tumor necrosis factor alpha-secreting, apoptosis-inducing macrophages than the tumors in the corresponding beta3 KO animals. The tumor-suppressive effects of host beta3 expression could be reversed by macrophage depletion or by transplantation of bone marrow from beta3 KO animals into beta3 WT animals, both of which significantly increased tumor growth independently of tumor vessel density. Taken together, these results show that host alphaVbeta3 integrin expression has opposing actions in the intracranial setting, enhancing tumor vascularization and growth while independently enhancing macrophage-mediated tumor elimination. Appropriate management of these functions could lead to enhanced efficacy of anti-integrin based therapies for glioma.     

Localization of 25-hydroxyvitamin D3-1 alpha-hydroxylase activity in the mammalian kidney

The tRNA from Ehrlich ascites tumor cells is deficient in the modified nucleoside Q (queuosine). Continuous infusion of Q base (queuine) to tumor-bearing mice reverses the deficiency of Q in Ehrlich ascites tRNA, and coincidently, causes an inhibition of tumor growth.     

APOPTOSIS OF EHRLICH MOUSE ASCITES CELLS INDUCED BY LONG-TERM EXPOSURE OF WEAK ELECTROMAGNETIC FIELDS IN VIVO

To study the possibility of apoptosis of tumor cells induced by weak electromagnetic fields (EMFs) in vivo, mice were inoculated with Ehrlich ascites cells and exposed to a long-term electromagnetic field (1 mT, 700 KHz). During the treatment, growth curves of mice were measured and compared between exposed and sham-exposed mice. The results show that the growth curves of healthy controls agree well with the ideal curve of logistic growth, but the growth curves of cancer mice deviate from the ideal curve. There is no difference in growth curves between exposed, and sham-exposed healthy mice, and they both agree with the ideal curve. However, a notable difference in growth curves between exposed and sham-exposed cancer mice was obtained. Moreover, the curves of sham-exposed mice deviate even more than those of the exposed mice; in other words, the growth curves of Ehrlich ascites mice deviate from the ideal curve of healthy mice but are shifted toward it by the EMF treatments. After the treatment, apoptosis of Ehrlich ascites cells from inoculated mice was analyzed by several methods, including flow cytometry, fluorescence microscopy, and DNA gel electrophoresis. Statistical analysis from flow cytometry shows that the apoptotic ratio of cells from exposed Ehrlich ascites mice was significantly higher than that from sham-exposed treated mice. Microscopic observation of Ehrlich ascites cells stained with acridine orange (AO) and propidium iodide (PI) showed typical apoptotic changes in exposed animals whose cell nuclei were highly condensed or fragmented and uniformly stained green by the AO, whereas cell nuclei from sham-exposed mice were stained green and showed a fine reticular pattern. Agarose gel electrophoresis of DNA from exposed mice showed that the chromatin DNA exhibited ladders, a characteristic feature of internucleosomal degradation of DNA by EMF treatments. For interactions between external electromagnetic fields and DNA, the mechanism of apoptosis of tumor cells induced by weak EMFs is discussed.     

Antitumor effect of lysine-isopeptides

Isopeptides (ε-peptides) of lysine, with a given Mw and low polydispersity (10–400 units), were synthesized to study the relationship between their chemical structure and biological effect. The designed compounds were of high purity, low polydispersity and high stereochemical purity. The effect of the compounds was tested on a human erythroleukemia cell line (K-562) and on four transplantable mouse tumors (L1210 lymphoid leukemia, P38 macrophage derived tumor, Ehrlich ascites carcinoma, Lewis lung tumor /LLT/). In case of the L1210 and P388 tumors and the Ehrlich carcinoma, survival of the animals was used as an indicator of the effect. In case of the Lewis lung tumor, the number and size of metastases in the lung and/or liver of treated and untreated mice were used as indicators. The polymers of polymerisation degree 80–120 (Mw 10.2–15.4 KD) showed the strongest antiproliferative effect both on K562 cells and the tumors growing in vivo. This effect was manifest with a significantly higher survival rate as compared to the control (L1210, P38, Ehrlich ascites), furthermore, by a decrease in the number and size of liver and lung metastases (LLT).     

N-Glycans differentially regulate eosinophil and neutrophil recruitment during allergic airway inflammation

Allergic airway inflammation, including asthma, is usually characterized by the predominant recruitment of eosinophils. However, neutrophilia is also prominent during severe exacerbations. Cell surface-expressed glycans play a role in leukocyte trafficking and recruitment during inflammation. Here, the involvement of UDP-N-acetylglucosamine:α-6-D-mannoside β1,6-N-acetylglucosaminyltransferase V (MGAT5)-modified N-glycans in eosinophil and neutrophil recruitment during allergic airway inflammation was investigated. Allergen-challenged Mgat5-deficient (Mgat5(-/-)) mice exhibited significantly attenuated airway eosinophilia and inflammation (decreased Th2 cytokines, mucus production) compared with WT counterparts, attributable to decreased rolling, adhesion, and survival of Mgat5(-/-) eosinophils. Interestingly, allergen-challenged Mgat5(-/-) mice developed airway neutrophilia and increased airway reactivity with persistent elevated levels of proinflammatory cytokines (IL-17A, TNFα, IFNγ)). This increased neutrophil recruitment was also observed in LPS- and thioglycollate (TG)-induced inflammation in Mgat5(-/-) mice. Furthermore, there was significantly increased recruitment of infused Mgat5(-/-) neutrophils compared with WT neutrophils in the peritoneal cavity of TG-exposed WT mice. Mgat5(-/-) neutrophils demonstrated enhanced adhesion to P-selectin as well as increased migration toward keratinocyte-derived chemokine compared with WT neutrophils in vitro along with increased calcium mobilization upon activation and expression of elevated levels of CXCR2, which may contribute to the increased neutrophil recruitment. These data indicate an important role for MGAT5-modified N-glycans in differential regulation of eosinophil and neutrophil recruitment during allergic airway inflammation.     

In vivo growth inhibitory and anti-angiogenic effects of synthetic novel dienone cyclopropoxy curcumin analogs on mouse Ehrlich ascites tumor

In the present study, four novel dienone cyclopropoxy curcumin analogs 1a–4a were synthesized by nucleophillic substitution reaction with cyclopropyl bromide. The tumor inhibitory and anti-angiogenic effects of the synthetic compounds were studied on mouse Ehrlich ascites tumor (EAT) in vivo. The compounds 1a–4a increased the life span (% ILS) of EAT bearing mice with corresponding significant reduction in ascites volume and cell number and induced apoptotic bodies in EAT cells. Anti-angiogenic studies of the compounds demonstrated significant reduction of microvessel density (MVD) in the peritoneum wall sections of mice and induced avascular zone in CAM model. Our findings demonstrate that the tumor growth inhibitory effects of synthetic dienone cyclopropoxy curcumin analogs 1a–4a could be mediated by promoting apoptosis and inhibiting tumor angiogenesis. However, the compounds need to be explored further to assess its clinical relevance.     

The problem of enhancing the biological action of ionizing radiations. Glucose as an agent for increasing the radiosensitivity of Ehrlich ascitic carcinoma cells in vitro

The effect of short-term incubation of Ehrlich ascites tumor cells in a medium containing excess glucose on their radiosensitivity was studied with a reference to the growth of tumors of ascitic and solid forms. It was shown that the incubation of cells with glucose, being accompanied by a change in pH of the suspension, caused, by itself, only a slight increase in the duration of the latent period of solid tumor formation and in the life-span of mice with ascitic tumors but increased considerably the lethal effect of radiation, as was estimated by the above mentioned criteria and the percentage of inoculated tumors.     

Targeted disruption of endothelial cell-selective adhesion molecule inhibits angiogenic processes in vitro and in vivo

Endothelial cell-selective adhesion molecule (ESAM) is a member of the immunoglobulin receptor family that mediates homophilic interactions between endothelial cells. To address potential in vivo angiogenic functions of this molecule, mice lacking ESAM (ESAM-/-) were generated by gene-targeted deletion. ESAM-/- mice did not show overt morphological defects in the vasculature. To evaluate the role of ESAM in pathological angiogenesis, wild type (WT) and ESAM-/- mice were injected with melanoma and Lewis lung carcinoma cells. By 14 days after injection, tumor volumes of B16F10 and LL/2 in ESAM-/- mice were 48 and 37% smaller, respectively, compared with WT mice. Vascular density of the tumors, as determined by CD31 staining, was also decreased in the ESAM null animals. Matrigel plug assays showed less neovascularization in ESAM-/- mice than in WT mice. ESAM-/- endothelial cells exhibited less in vitro tube formation and decreased migration in response to basic fibroblast growth factor when compared with WT cells, and endothelial-like yolk sac cells engineered to overexpress ESAM showed accelerated tube formation in vitro. These in vitro and in vivo studies suggest that ESAM has a redundant functional role in physiological angiogenesis but serves a unique and essential role in pathological angiogenic processes such as tumor growth.     

Characterization of LPS-induced lung inflammation in cftr-/- mice and the effect of docosahexaenoic acid.

The mechanism by which Pseudomonas causes excessive inflammation in the cystic fibrosis lung is unclear. We have reported that arachidonic acid is increased and docosahexaenoic acid (DHA) decreased in lung, pancreas, and ileum from cftr-/- mice. Oral DHA corrected this defect and reversed the pathology. To determine which mediators regulate inflammation in lungs from cftr-/- mice and whether inhibition occurs with DHA, cftr-/- and wild-type (WT) mice were exposed to aerosolized Pseudomonas lipopolysaccharide (LPS). After 2 days of LPS, tumor necrosis factor-alpha (TNF-alpha), macrophage inflammatory protein-2, and KC levels in bronchoalveolar lavage fluid were increased in cftr-/- compared with WT mice and not suppressed by pretreatment with oral DHA. Neutrophil levels were not different between cftr-/- and WT mice. After 3 days of aerosolized LPS, neutrophil concentration, TNF-alpha, and the eicosanoids 6-keto-PGF1alpha, PGF2alpha, PGE2, and thromboxane B2 were all increased in bronchoalveolar lavage fluid from cftr-/- mice compared with WT controls. Oral DHA had no significant effect on TNF-alpha levels in cftr-/- mice. In contrast, neutrophils and eicosanoids were decreased in cftr-/- but not in WT mice treated with DHA, indicating that the effects of DHA on these inflammatory parameters may be related to correction of the membrane lipid defect.     

Antigenicity and Immunogenicity of Cell Surface Fraction of Ehrlich Ascites Tumor Cells

A membrane fraction corresponding to one-tenth of the dry weight of the whole cell was prepared from Ehrlich ascites tumor cells. Such a fraction retained antigenicity in immune adherence inhibition test using antisera taken from mice hyperimmunized to Ehrlich tumors. The antigenicity was impaired by freezing and thawing, and disappeared after such treatment was repeated more than five times, but was stable to heating up to 60°. The above fraction retained immunogenicity to induce resistance of mice against Ehrlich tumors and retained considerable immunogenicity after drying with acetone-ether.     

中药麝香胶囊抑瘤作用的实验研究

目的探讨中药麝香胶囊对小鼠实验性肿瘤的疗效。方法昆明小鼠分别接种艾氏腹水癌、S-180、肝癌细胞株（hepatocellular carcinoma,HCC）三种癌细胞株,接种24h后开始给予中药麝香胶囊,每日1次,共10d。分高、中、低三个剂量给药（4、2、1g／kg以主药麝香药量计）。阳性对照以天仙丸胶囊（1g／kg）一次性灌服。阴性对照组以同体积生理盐水灌服。接种实体瘤动物于第10天处死,称瘤重,计算抑瘤率;接种腹水瘤动物,观察存活时间,计算生命延长率。结果中药麝香胶囊对实体癌有一定的抑瘤作用,但未达到药典规定的抑瘤率30％的要求;对腹水癌也有一定的抑瘤作用,但也未达到药典规定生命延长率50％的要求;统计学分析（P〉0．05）差异没有显著性。结论中药麝香胶囊抑瘤作用不明显,其配方及剂型有待进一步研究与改进。     

Monocytes are potent facilitators of alveolar neutrophil emigration during lung inflammation: role of the CCL2-CCR2 axis

Coordinated neutrophil and monocyte recruitment is a characteristic feature of acute lung inflammatory responses. We investigated the role of monocyte chemotactic protein-1 (CCL2, JE) and the chemokine receptor CCR2 in regulating alveolar leukocyte traffic. Groups of wild-type (WT) mice, CCR2-deficient mice, lethally irradiated CCR2-deficient and WT mice that were reciprocally bone marrow transplanted (chimeric CCR2 deficient and WT, respectively), chimeric CCR2-deficient mice with an enriched CCR2(+) alveolar macrophage population, and CCR2-deficient mice transfused with CCR2(+) mononuclear cells were treated with intratracheal CCL2 and/or Escherichia coli endotoxin. Our data show that alveolar monocyte recruitment is strictly dependent on CCR2. LPS-induced neutrophil migration to the lungs is CCR2 independent. However, when CCR2-bearing blood monocytes are present, alveolar neutrophil accumulation is accelerated and drastically amplified. We suggest that this hitherto unrecognized cooperativity between monocytes and neutrophils contributes to the strong, coordinated leukocyte efflux in lung inflammation.     

Effect of centimeter microwaves and the combined magnetic field on the tumor necrosis factor production in cells of mice with experimental tumors

The effect of fractionated exposure to low-intensity microwaves (8.15-18 GHz, 1 microW/cm2, 1.5 h daily for 7 days) and combined weak magnetic field (constant 65 1 microT; alternating--100 nT, 3-10 Hz) on the production of tumor necrosis factor in macrophages of mice with experimental solid tumors produced by transplantation of Ehrlich ascites carcinoma was studied. It was found that exposure of mice to both microwaves and magnetic field enhanced the adaptive response of the organism to the onset of tumor growth: the production of tumor necrosis factor in peritoneal macrophages of tumor-bearing mice was higher than in unexposed mice.