Effects of retinoic acid, auranofin and mercuric chloride on plasminogen activator activity in post-implantation cultured mouse embryos

Plasminogen activator (PA) activity has been suggested to be an important determinant of cell migration and tissue modelling during organogenesis. We have postulated that in the developing embryo, any abnormal modulation of this enzymatic activity may lead to the production of teratogenic effects. In the present study, we investigated the effect of teratogenic doses (inducing about 50% of malformed embryos) of retinoic acid, auranofin and mercuric chloride on PA activity in post-implantation cultured mouse embryos. At the end of the culture period, PA activities of malformed and normal embryos in the same treatment group were compared. PA activities in compound-exposed embryos were also compared with those in untreated controls. The design of the present experiment allowed the identification of the effect of drugs on PA activity and its possible relation with induced malformations. An increased PA activity was observed in malformed embryos treated with mercuric chloride. PA activity was slightly increased in both groups (normal and malformed) exposed to retinoic acid. No effect of auranofin was observed on embryo PA activity. In conclusion, the data do not confirm but they also do not contradict the hypothesis that abnormal levels of PA activity lead to dysmorphogenesis.     

In vitro effect of mercury and vanadium on superoxide anion production and plasminogen activator activity of mouse peritoneal macrophages

The in vitro effects of mercuric chloride and vanadate were examined on two functions of mouse peritoneal macrophages, i.e., the superoxide anion production and the plasminogen activator (PA) activity. Vanadate, at concentrations which do not affect the viability of the cells, does not seriously alter any of these functions. High concentrations of mercury depress the respiratory burst; this effect results from loss of the reducing properties of cellular NADPH. Low concentrations of mercury stimulate the effect of phorbol 12-myristate 13-acetate on PA activity. The mechanism of this stimulation does not involve the protein kinase C system. It is hypothesized that mercury could enhance the synthesis of PA, its translocation to the cell surface, or its binding to the membrane receptors.     

绿原酸对Lewis肺癌小鼠及人A549肺癌的实验研究

目的 研究绿原酸对肺癌的影响及联合用药后对化疗药物的增敏作用.方法 采用MTT法研究绿原酸对人肺癌A549细胞增殖的影响,绿原酸对移植瘤的影响及对化疗药物的增敏作用.结果 绿原酸对人肺癌A549细胞增殖无影响,对Docetaxel(1.6 μg·mL-1)无增敏作用;对A549肺癌有一定抑制趋势,可明显抑制Lewis肺癌的生长;联合用药在一定程度上可提高抗肿瘤作用,但对化疗药物的抗肿瘤作用无协同性.结论 绿原酸对人肺癌A549细胞无细胞毒作用;对A549肺癌无明显抑制作用,但可明显抑制Lewis肺癌的生长,联合阿霉素用药对抗肿瘤无增敏作用.     

Antitumor activity of ascorbic acid in combination with antitumor agents against Lewis lung carcinoma

Antitumor activity of ascorbic acid in combination with antitumor agents (mitomycin C and 5-fluorouracil) was examined against subcutaneously implanted Lewis lung carcinoma-bearing C57BL/6 mice by feeding them an ascorbic acid-deficient diet. The mice were divided into four groups: group 1 received intraperitoneally mitomycin C (2 mg/kg) and 5-fluorouracil (50 mg/kg) once a week for four weeks beginning from the day after implantation of tumors, as well as ascorbic acid (1000 mg/kg) twice a week for the same four weeks; group 2 received only mitomycin C and 5-fluorouracil; group 3 received only ascorbic acid; group 4 received a vehicle (physiological saline). Tumor growth of group 1 compared with the other three groups, and that of group 2 compared with groups 3 and 4, was significantly inhibited by day 13 post implantation. Histological examinations of tumor tissues at 10 days after implantation of tumors already showed degenerative changes which indicated these antitumor effects.     

川芎嗪对小鼠Lewis肺癌的治疗作用

目的:研究中药川芎嗪(TMP)对实体肿瘤的疗效及其可能的作用机理。方法:本研究运用Lewis肺癌小鼠模型,观察TMP对小鼠Lewis肺癌的疗效、免疫功能、生存质量及不良反应的影响。结果:TMP治疗小鼠Lewis肺癌能抑制肿瘤的生长,稳定病灶,提高荷瘤小鼠的生存质量,改善免疫功能。结论:TMP对小鼠Lewis肺癌有明显疗效,其作用机理与其本身对Lewis肺癌实体瘤有抑制作用和提高荷瘤小鼠免疫功能有关。     

The actions of retinal and retinoic acid on histamine release from rat peritoneal mast cells

Histamine release from rat peritoneal mast cells induced by anti-IgE was potentiated by the retinoids: retinoic acid 2-10 microM and retinal 1-5 microM. Retinal also produced a concentration-dependent increase in anti-IgE-stimulated 45Ca uptake by these mast cells. A similar potentiating action of both retinoids was observed on histamine release induced by 12-O-tetradecanoylphorbol-13-acetate (TPA). For both anti-IgE- and TPA-induced histamine release, the potentiating effect of the two retinoids was only observed in the presence of extracellular calcium. In contrast, histamine release induced by compound 48/80 was inhibited by retinal 1-5 microM and by retinoic acid 10-50 microM and the inhibition was the same in the presence as in the absence of extracellular calcium 1 mM. Histamine release induced by calcium and the calcium ionophore A 23187 was inhibited by retinoid acid 2-10 microM and by retinal 10 microM. Inhibitions of compounds 48/80-induced histamine release by cromoglycate and by retinal were additive. It is concluded that while the actions of retinoids on rat peritoneal mast cells are consistent with the inhibition of protein kinase C, another action of these compounds, unrelated to this enzyme, may explain the data.     

Inhibition by auranofin of the production of prostaglandin E2 and nitric oxide in rat peritoneal macrophages

In rat peritoneal macrophages, 12-O-tetradecanoylphorbol 13-acetate (TPA) (16.2 nM) stimulated production of both prostaglandin E2 and nitric oxide. TPA also increased the levels of mRNA for cyclooxygenase-2 and inducible nitric oxide synthase, suggesting that the increase in the production of prostaglandin E2 and nitric oxide is due to the increase in the levels of mRNA for cyclooxygenase-2 and inducible nitric oxide synthase, respectively. The TPA-induced increase in prostaglandin E2 production was partially inhibited by the inhibitor of nitric oxide synthase L-N(G)-monomethyl-L-arginine acetate (L-NMMA), and the TPA-induced increase in nitric oxide production was partially inhibited by the cyclooxygenase inhibitor indomethacin, suggesting that both the production of prostaglandin E2 and nitric oxide in TPA-stimulated macrophages is influenced by each other. The orally active chrysotherapeutic agent auranofin, at 3 and 10 microM, inhibited the TPA-stimulated production of prostaglandin E2 and nitric oxide, and suppressed the TPA-induced increase in the levels of mRNA for cyclooxygenase-2 and inducible nitric oxide synthase. These findings indicate that the inhibition by auranofin of the TPA-stimulated production of prostaglandin E2 and nitric oxide is due to the decrease in the levels of mRNA for cyclooxygenase-2 and inducible nitric oxide synthase, respectively, and the interaction of the production between prostaglandin E2 and nitric oxide may partly be involved in the mechanism for the inhibition by auranofin of the production of both prostaglandin E2 and nitric oxide.     

CIK细胞与LAK细胞对Lewis肺癌小鼠的抑瘤作用

目的观察CIK(cytokinc induced Killer)细胞及LAK(1ymphokine activated killer)细胞对Lewis肺癌小鼠的抑瘤作用。方法建立Lewis肺癌小鼠动物模型，静脉输入CIK细胞做抗肿瘤治疗，同时设LAK对照，用同位素法检测经治疗后荷瘤鼠T细胞增殖及NK细胞毒活性。结果CIK细胞对Lewis肺癌小鼠的抗肿瘤作用显著强于LAK细胞。结论CIK细胞对Lewis肺癌小鼠有明显的抗肿瘤作用，并且优于LAK细胞。     

Influence of isomerisation on the growth inhibitory effects and cellular activity of 13-cis and all-trans retinoic acid in neuroblastoma cells

Treatment with 13-cis retinoic acid (13-cis RA) has been shown to significantly improve the clinical outcome of children with high-risk neuroblastoma. Despite the large number of studies investigating the cellular effects of retinoids in neuroblastoma cells, the influence of RA isomerisation and the factors that determine the extent of RA isomerisation and uptake are unknown. The aim of this study was to establish the extent of extra- and intracellular isomerisation of 13-cis RA and all-trans retinoic acid (ATRA) in neuroblastoma cell lines, and to investigate the influence of isomerisation on their growth inhibitory effects and on the regulation of expression of cellular retinoic acid binding protein II (CRABP II) and RAR-beta. Limited extracellular isomerisation was observed up to 72 hr after incubation of four neuroblastoma cell lines with 10 microM 13-cis RA or ATRA. The retinoic acid isomer present initially in the medium accounted for >75% of extracellular retinoid exposure. By contrast, incubation with 13-cis RA resulted in intracellular levels of ATRA comparable to those of 13-cis RA. This degree of intracellular isomerisation was not observed after ATRA incubations, with 13-cis RA accounting for <10% of total intracellular retinoids. No differences were observed in the sensitivity of three N-type neuroblastoma cell lines to either 13-cis RA (IC(50): 11.2-13.9 microM) or ATRA (IC(50): 12.9-14.4 microM), despite 10-fold differences in intracellular retinoid levels. A decrease in sensitivity to 13-cis RA (IC(50)=137 microM), as compared to ATRA (IC(50)=41 microM), was observed in the S-type cell line SH S EP. RAR-beta was induced in a dose-dependent manner in SH SY 5Y cells following incubation with ATRA, whereas a weaker and delayed induction was observed with 13-cis RA. Similarly, incubation with ATRA resulted in a greater induction of CRABP II in these cells. In summary, these results indicate either an intracellular conversion of 13-cis RA to ATRA or a selective uptake of ATRA and suggest that this may mediate the differential activity of 13-cis RA in neuroblastoma cell subtypes.     

Anti-tumorigenic activity of sophoflavescenol against Lewis lung carcinoma in vitro and in vivo

This study examined the in vitro cytotoxic activity and in vivo antitumor activity as well as intracellular apoptotic capacities of a prenylated flavonol, sophoflavescenol from Sophora flavescens, to evaluate prospective anti-tumorigenic drugs, and antitumor potential. In addition, the in vitro antioxidant and anti-inflammatory capacities were evaluated. Despite the small effect on human breast adenocarcinoma (MCF-7), sophoflavescenol showed cytotoxicity against human leukaemia (HL-60), Lewis lung carcinoma (LLC), and human lung adenocarcinoma epithelial (A549) cells. Interestingly, it also exerted potent in vivo antitumor activity by tumor growth inhibition in the LLC tumor model as well as apoptotic activity by caspase-3 activation in HL-60 cells. In addition, it exhibited potent antioxidant activities in 1,1-diphenyl-2-picrylhydrazyl, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt radicals and lipid peroxidation assays. Sophoflavescenol exerted notable anti-inflammatory activity by inhibiting nitric oxide generation and tert-butylhydroperoxide-induced ROS generation rather than inhibiting nuclear factor kappa B activation in RAW 264.7 cells. The findings show that the antioxidant, anti-inflammatory, and apoptotic activities of sophoflavescenol might contribute to the antitumor activity without severe side effects, highlighting its potential for chemoprevention and/or anticancer drugs due to multi-effective targets in almost all stages of tumorigenesis, including initiation, promotion, and progression.     

Antitumoral effects of Pseudomonas aeruginosa lectins on Lewis lung carcinoma cells cultured in vitro without and with murine splenocytes.

Examination of the in vitro effects of PA-I and PA-II lectins of Pseudomonas aeruginosa on Lewis lung carcinoma cells revealed that these lectins differ in their effects. PA-I, the galactophilic lectin, exhibited both cytotoxic and cytostatic activities on these cells (tested by [3H]thymidine incorporation and by crystal violet vital staining). The two activities were dose and time dependent and inhibitable by the addition of methyl-alpha-D-galactoside to the culture medium. PA-II, the L-fucose and D-mannose binding lectin of the same Pseudomonas strain did not exhibit such a direct toxic effect on the tumor cells but affected them in the presence of splenocytes. Its addition to the tumor cells cocultured with murine (C57B1) splenocytes led to a profound cytolysis of the tumor cells, an effect which was inhibited by L-fucose.     

Comparison of the effects of multiwall carbon nanotubes on the epithelial cells and macrophages

Abstract

Effects of two kinds of multiwall carbon nanotubes (MWCNTs) on cells were examined. The effects of MWNT-7, which has been reported to be carcinogenic, and MWCNT-B, whose toxicity is unclear, were examined in both epithelial cells and macrophages. Human lung carcinoma A549 cells were used as representative epithelial cells and differentiated human monocyte THP-1 cells, as well as rat pulmonary macrophages NR8383, were employed to examine possible harmful effects of the MWCNTs. The MWCNTs induced the production of chemokines such as interleukin-8 (IL-8). MWCNTs were found to more strongly affect macrophages than epithelial cells. In addition, the toxicity was more pronounced in the MWNT-7 exposed cells than in those exposed to MWCNT-B. Cytochalasin D and amiloride treatment of differentiated THP-1 cells reduced cell-associated MWCNTs and IL-8 induction. To confirm these cellular influences in vivo, intratracheal administration of each type of MWCNT was performed by pharyngeal aspiration in the mouse lung. Analysis of bronchoalveolar lavage fluid (BALF) showed increase of inflammatory monocyte in MWNT-7 exposed animals at 1week after. In addition, neutrophils in the BALF were also significantly increased MWNT-7 exposed animals at 1 week and 1 month after. Aspiration of MWNT-7 caused formation of granulomas in the lung. Formation of the granulomas was not observed in the case of MWCNT-B. These results suggest that cellular uptake of the MWCNTs by phagocytosis and chemokine induction is important aspects of their toxicity.     

塞来昔布联合吉西他滨对Lewis肺癌皮下移植瘤的抑制作用

目的研究塞来昔布(CEL)与吉西他滨(GEM)联用对Lewis肺癌皮下移植瘤的抑制效应。方法 40只小鼠随机分为对照组(A组)、GEM组(B组)、CEL组(C组)和CEL/GEM联用组(D组),每组10只。A、B组给予普通饲料;C、D组自接种后第2天给予含CEL 1mg/g的饲料喂养。B、D组于肿瘤接种后第7天,给予GEM 100mg/kg溶液0.2ml腹腔注射;A、C组给予等量生理盐水。每周注射1次,连续3周。实验期间测量肿瘤长短径,并绘制肿瘤生长曲线,4周后处死,称量瘤重,计算抑瘤率;检测移植瘤组织血管内皮生长因子(VEGF)表达、微血管密度(MVD)和肿瘤细胞的凋亡。结果 B、C和D组抑瘤率分别为54.18%、49.52%和88.59%;D组移植瘤组织的VEGF表达及MVD低于A、B和C组(P<0.05),而凋亡指数高于A、B和C组(P<0.05)。结论 CEL和GEM均能抑制Lewis肺癌移植瘤的生长,两者联用通过抑制肿瘤微血管生成及诱导肿瘤细胞凋亡发挥协同抗肿瘤效应。     

Anti-tumor and anti-angiogenic effects of Phyllanthus urinaria in mice bearing Lewis lung carcinoma

Phyllanthus urinaria, a widely used herb medicine in Asia, was tested for its anti-tumor effect in vivo for the first time. The anti-tumor activity in P. urinaria extract was evaluated by its effect on tumor developed in C57BL/6J mice with implantation of Lewis lung carcinoma cells. The oral administration of P. urinaria to mice caused significant inhibition of tumor development with lower occurrence rate and markedly reduced tumor size. Neither the total body weight of mouse nor the weights of organs including heart, lung, liver, spleen and kidney revealed any difference between two groups, suggesting limited in vivo cytotoxic effect of P. urinaria in mice. TUNEL assay demonstrated the increase of apoptosis in tumor sections prepared from P. urinaria-treated mice compared with control mice. It is worth of note that the neovascularization in tumor was inhibited in P. urinaria-treated mice, which implicated the potential anti-angiogenic effect of P. urinaria. Further study using an in vitro matrix-induced tube formation of HUVECs again confirmed the anti-angiogenic action of P. urinaria. P. urinaria exerted no inhibitory effect on the growth of HUVECs, however, the migration of HUVECs as analyzed using transwell assay was suppressed markedly by P. urinaria in a dose-dependent manner. All together, the present study indicated that P. urinaria extract is an anti-tumor and anti-angiogenic agent, which can be used safely in animals.     

Plasminogen activator activity of cultured murine macrophages and effects of isopropylmethylphosphonofluoridate (sarin)

Casein-elicited mouse peritoneal macrophages cultured in the presence of phorbol 12-myristate 13-acetate (PMA) express u-PA. Induction is maximal after 4 hr of stimulation and u-PA activity is mainly recovered with the membrane fraction of the cellular lysate. This enzymatic activity is inhibited by isopropylmethylphosphonofluoridate after stimulation by PMA. The presence of the organophosphorus compound before stimulation does not affect the activity. The results of the present study on the kinetics of u-PA activity in cultured macrophages, its subcellular localization and the effect of an organophosphorus ester are consistent with the concept that the development of pericellular proteolysis proceeds through a series of stages, namely, (a) synthesis of pro-u-PA, (b) binding to membrane receptors, (c) activation to a double-chain u-PA, and (d) conversion of plasminogen into plasmin. The assay procedure developed here provides a sensitive tool to investigate the mechanism of interference of chemicals with several steps of induction of this enzymatic activity in macrophages.     

Inhibitors of the proteolytic activity of urokinase type plasminogen activator

Urokinase type plasminogen activator (uPA) activates plasminogen to plasmin and is often associated with diseases where tissue remodeling is essential (e.g. cancer, macular degeneration, atherosclerosis). We discuss some of the mechanisms of uPA action in diseases, and evidence that some of the early uPA inhibitors can modulate the progression of these diseases. Recently, a number of research groups have discovered, with the aid of structure-based design, a new generation of uPA inhibitors. These inhibitors are much more potent and selective than their predecessors. We will review this progress here, and give particular attention to the structural rationale associated with these observed increases in potency and selectivity.     

Plasminogen activator activity of normal and retinoic acid-treated post-implantation embryos

Urokinase-type plasminogen activator (uPA) has been implicated in cellular migration accompanying different biological phenomena including organogenesis. An increase in uPA activity was observed in mouse post-implantation embryos during the early organogenesis period. Since we have previously shown that all-trans retinoic acid (RA) prevented the induction of uPA in mouse peritoneal macrophages, we have now assessed whether teratogenic doses of this agent could also interfere with uPA activity in mouse embryo in vitro and in vivo. Post-implantation embryos (8.5 days) were incubated for up to 24 hr with micromolar concentration of RA resulting in the occurrence of malformations. No significant difference in uPA activity was found between control and treated embryos. Likewise, uPA activity was not altered in embryos explanted on day 9.5 from dams treated 24 hr before with a teratogenic dose of RA. This study indicates that the teratogenic activity of RA is not caused by an inhibition of the induction of uPA in embryos.     

The long-term effects of metoprolol and epanolol on tissue-type plasminogen activator and plasminogen activator inhibitor 1 in patients with ischaemic heart disease

This double-blind, randomized parallel group study investigated the effect of 6 months -adrenoceptor antagonist therapy with either metoprolol (₁-selective without intrinsic sympathomimetic activity [ISA]) or epanolol (₁-selective with ISA) on markers of endogenous fibrinolysis in 20 patients with chronic stable angina receiving concurrent treatment with nifedipine.Neither drug had an effect on tissue-type plasminogen activator or plasminogen activator inhibitor type 1 (PAI-1). A significant correlation between fasting insulin and PAI-1 has previously been described and was confirmed in this study. The group treated with metoprolol showed a significant rise in fasting insulin after 6 months with no change in PAI-1.This suggests that the previously described link between these two may not be causal.     

SP-40,40 is a component of plasminogen activator inhibitor-1-binding protein and stabilizes plasminogen activator inhibitor-1 activity

A complex of plasminogen activator inhibitor-1 (PAI-1) and PAI-1-binding protein (PAI-1-BP) contained S-protein (vitronectin), PAI-1 and unidentified 40-kDa protein on SDS-PAGE under reducing conditions. By Western-blot analysis, the 40-kDa protein was identified as SP-40,40 using anti-SP-40,40 antibody. Therefore, it was thought that PAI-1-BP consisted of S-protein and SP-40,40. It is known that PAI-1 is a labile protein which becomes inactive during incubation at 37 degrees C. However, after the incubation of PAI-1 with SP-40,40 at 37 degrees C for 1 h, PAI-1 could still form a complex with tissue plasminogen activator (tPA), and it inhibited plasmin formation in the mixture of plasminogen and urine plasminogen activator (uPA). The results clearly indicated that SP-40,40 stabilized PAI-1 activity as well as S-protein did.