Suppression of <i>N</i>-Methyl-<i>N</i>-Nitrosourea-Induced Retinal Damage in Mice by Oligonol, an Oligomerized Polyphenol Formulation

Oligonol is a lychee fruit-derived functional food that contains oligomerized polyphenol compounds. Oligonol exhibits a number of beneficial biological effects, primarily due to its antioxidant activity. Retinitis pigmentosa (RP) is an inherited chronic degenerative disease affecting retinal photoreceptor cells. There is currently no effective therapy capable of stopping or reversing the progression of the disease. In RP, apoptosis of photoreceptor cells resulting from oxidative damage is considered to be the final common pathway. In this report, we present an evaluation of the suppressive activity of Oligonol against N-methyl-N-nitrosourea (MNU)-induced retinal damage in mice, which is a commonly used animal model of RP. Both intraperitoneal and oral administration of Oligonol reduced the loss of photoreceptor cells 7 days after MNU injection, as evaluated by histological staining. Photoreceptor cells derived from MNU-treated mice exhibited increased TUNEL-positive staining, suggesting increased DNA fragmentation, a hallmark of apoptosis. Oligonol treatment reduced the number of TUNEL-positive cells. Additionally, Oligonol suppressed MNU-induced retinal production of 8-hydroxydeoxyguanosine (8-OHdG), a marker of oxidative stress. Moreover, Oligonol attenuated the MNU-induced decrease in the visual activity of mice, as evaluated by the visual cliff test. Oligonol, therefore, effectively suppresses NMU-induced retinal degeneration.

Protective effects of naringenin eye drops on N-methylN-nitrosourea-induced photoreceptor cell death in rats

AIM:To investigate the effects of naringenin eye drops on N-methyl-N-nitrosourea (MNU)-induced photoreceptor cell death in rats.METHODS:Photoreceptor cell death was induced by single intraperitoneal injection of MNU(60 mg/kg)in rats.Both eyes of all animals were instilled with one drop of vehicle,0.5% or 1.0% naringenin eye drops three times per day from 7d before to 17d after MNU injection.Effects of naringenin on MNU-induced photoreceptor cell death were evaluated by electrophysiological and histological analysis.RESULTS:Flash electroretinography (FERG)and oscillatory potentials (OPs) recordings showed that the vehicle control group had remarkable reduction of amplitudes and prolongation of latency times.FERG and OPs responses were significantly reversed in MNUinduced rats treated with 0.5%or 1.0% naringenin eye drops compared with the vehicle control.The retinal morphological results showed that naringenin dosedependently preserved the outer nuclear layer,outer retina and total retina.CONCLUSION:These results indicate that topical treatment with naringenin eye drops prevented retinal neurons from MNU-induced structural and functional damages.

1,3-Bis(2-chloroethyl)-1-nitrosourea enhances the inhibitory effect of Resveratrol on 5-fluorouracil sensitive/resistant colon cancer cells

AIM:To study the mechanism of 5-fluorouracil(5-FU)resistance in colon cancer cells and to develop strategies for overcoming such resistance by combination treatment.METHODS:We established and characterized a 5-FU resistance(5-FU-R)cell line derived from continuous exposure(25μmol/L)to 5-FU for 20 wk in 5-FU sensitive HCT-116 cells.The proliferation and expression of different representative apoptosis and anti-apoptosis markers in 5-FU sensitive and 5-FU resistance cells were measured by the MTT assay and by Western blotting,respectively,after treatment with Resveratrol(Res)and/or 1,3-Bis(2-chloroethyl)-1-nitrosourea(BCNU).Apoptosis and cell cycle arrest was measured by 4',6'-diamidino-2-phenylindole hydrochloride staining and fluorescence-activated cell sorting analysis,respectively.The extent of DNA damage was measured by the Comet assay.We measured the visible changes in the DNA damage/repair cascade by Western blotting.RESULTS:The widely used chemotherapeutic agents BCNU and Res decreased the growth of 5-FU sensitive HCT-116 cells in a dose dependent manner.Combined application of BCNU and Res caused more apoptosis in5-FU sensitive cells in comparison to individual treatment.In addition,the combined application of BCNU and Res caused a significant decrease of major DNA base excision repair components in 5-FU sensitive cells.We established a 5-FU resistance cell line(5-FU-R)from 5-FU-sensitive HCT-116(mismatch repair deficient)cells that was not resistant to other chemotherapeutic agents(e.g.,BCNU,Res)except 5-FU.The 5-FU resistance of 5-FU-R cells was assessed by exposure to increasing concentrations of 5-FU followed by the MTT assay.There was no significant cell death noted in5-FU-R cells in comparison to 5-FU sensitive cells after5-FU treatment.This resistant cell line overexpressed anti-apoptotic[e.g.,AKT,nuclear factorκB,FLICE-like inhibitory protein),DNA repair(e.g.,DNA polymerase beta(POL-β),DNA polymerase eta(POLH),protein Flap endonuclease 1(FEN1),DNA damage-binding protein 2(DDB2)]and 5-FU-resistance proteins(thymidylate synthase)but under expressed pro-apoptotic proteins(e.g.,DAB2,CK1)in comparison to the parental cells.Increased genotoxicity and apoptosis were observed in resistant cells after combined application of BCNU and Res in comparison to untreated or parental cells.BCNU increased the sensitivity to Res of 5-FU resistant cells compared with parental cells.Fifty percent cell death were noted in parental cells when 18μmol/L of Res was associated with fixed concentration(20μmol/L)of BCNU,but a much lower concentration of Res(8μmol/L)was needed to achieve the same effect in 5-FU resistant cells.Interestingly,increased levels of adenomatous polyposis coli and decreased levels POL-β,POLH,FEN1 and DDB2 were noted after the same combined treatment in resistant cells.CONCLUSION:BCNU combined with Res exerts a synergistic effect that may prove useful for the treatment of colon cancer and to overcome drug resistance.

Meclizine Chloridrate and Methyl-β-Cyclodextrin Associated with Monophosphoester Synthetic Phosphoethanolamine Modulating Proliferative Potential in Triple-Negative Breast Cancer Cells

Synthetic phosphoethanolamine(Pho-s)is a monophosphoester ester with anti-inflammatory and pro-apoptotic properties.Meclizine chloridrate(MC)is a histamine H1 receptor blocker that is also able to inhibit cellular respiration.However,MC does not inhibit cellular respiration in isolated mitochondria such as antimycin and rotenone.Methyl-β-cyclodextrin(MβCD)belongs to theβ-cyclodextrin family,which is capable of removing cholesterol from the plasma membrane.The aim of this study was to evaluate the proliferative effects of meclizine chloridrate and methyl-β-cyclodextrin compounds associated with synthetic phosphoethanolamine in a triple-negative human breast tumor line,MDA-MB-231 Cell viability of the tumor line and normal cells FN1 was evaluated by MTT colorimetric test;the production of free radicals was determined by lipoperoxidation(LPO)test;and the percentage of cell cycle phases and proliferative index was evaluated by flow cytometry.Cell viability demonstrated a significant decrease with the treatments of MβCD,MC and Pho-s associated with MC.The production of free radicals decreases significantly in all treatments.In addition,a significant increase of DNA fragment and decrease in G0/G1 cell cycle phase were observed in cellular percentage with concentrations of 20 and 30 mM of Pho-s in association with MC and MβCD,respectively.

Auxiliary Ligands Mediated One- and Two-dimensional Cd(Ⅱ) Coordination Polymers Incorporating Methyl-3- hydroxy-5-carboxy-2-Thiophenecarboxylate Ligand

Two N-donor ligands mediated Cd(II) coordination polymers, namely, [Cd(L)(dmpz)_2]_n(1) and {[Cd(L)(atr)_(0.5)(H_2O)](H_2O)}n(_2)(H_2 L = methyl-3-hydroxy-5-carboxy-2-thiophenecarboxylate, dmpz = 3,5-dimethylpyrazole, and atr = 4-amino-1,2,4-triazole), have been produced. Their structures were characterized by single-crystal X-ray diffraction analysis, elemental analyses and infrared spectra. Compound 1 possesses a one-dimensional(1 D) chain structure and is finally extended into a three-dimensional(3 D) supramolecular architecture though hydrogen bonding interactions. Compound 2 features a two-dimensional(2 D) network with 4-connected sql topology based on dinuclear Cd(II) clusters as nodes, which is also assembled into a 3 D supramolecular architecture through hydrogen bonding interactions. Furthermore, compounds 1 and 2 exhibit high thermal stabilities and intense fluorescent emission in the solid, and can be explored as potential luminescent materials.

SYNTHESES OF N- METHYL-^(14)C BENZYL AMINE AND N- METHYL-^(14)C BENZYL NITROSAMINE

The syntheses of N-Methyl-14C benzyl amine and N-Methyl-14C benzyl nitrosamine are reported. Specific activities were approximately 920 MBq/mmol. Chemical and radiochemical purity checked by HPLC were more than 95%.

In vitro release of 1,3-bis(2-chloroethyl)-1-nitrosourea sustained-release microspheres and the distribution in rat brain tissues

BACKGROUND: The implantation of released chemotherapeutic drugs, which takes biodegradable polymer as vector, into the tumor site can get high concentration and release the drug for a long time, it can directly act on the tumor cells, and reduce the general toxicity. OBJECTIVE: To explore the in vitro and in vivo course of 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) sustained-release from BCNU-loaded polylactide (PLA) microspheres (MS) and location in rat brain tissue. DESIGN: A repetitive measurement. SETTING:Central Pharmacy, General Hospital of Chinese People's Armed Police Forces. MATERIALS: Thirty male SD rats were used. PLA (Mr5000, batch number: KSL8377) was produced by Wako Pure Chemical Inc.,Ltd. (Japan); BCNU (batch number: 021121) by Tianjin Jinyao Amino Acid Co., Ltd.; BCNU-PLA-MS was prepared by the method of solvent evaporation and pressed into tablets (10 mg/tablet). High-performance liquid chromatography (HPLC) Agilent 1100 (USA); LS9800 liquid-scintillation radiometric apparatus (Beckman). Chromatographic conditions: Elite Hypersil ODS2 C18 chromatographic column (5 μm, 4.6 mm×150 mm); Mobile phase: methanol: water (50:50), flow rate was 1.0 mL per minute, wave length of ultraviolet detection was 237 nm, and the inlet amount of samples was 10 μL. METHODS: The experiments were carried out in the experimental animal center of the General Hospital of Chinese Armed Police from May 2004 to July 2005. ① In vitro BCNU-PLA-MS release test: BCNU-PLA-MS was prepared by the method of solvent evaporation, then placed in 0.1 mol/L phosphate buffered solution (PBS, pH 7.4, 37 ℃), part of MS were taken out at 1, 2, 3, 7, 10 and 15 days respectively, and the rest amount of BCNU in MS was determined by HPLC, then the curve of BCNU-PLA-MS release was drawn. ②In vivo BCNU-PLA-MS release and distribution test: The rats were anesthetized, then BCNU-PLA-MS were implanted to the site 1 mm inferior to the cortex of frontal lobe. Five rats were killed postoperatively at 4 hours, 1, 2, 3, 7 and 15 days, the residual MS was removed from the brain tissue. The rest amount of BCNU was determined with HLPC, and the curve of BCNU-PLA-MS release was drawn as compared with the amount of BCNU in the implanted tablets. Besides, brain tissues (1 g) at the implanted side and the contralateral one were obtained respectively, blood sample (0.5 mL) was also collected, 3H-BCNU was counted radioactively in radioactive liquid flash solution. The distributions of BCNU-PLA-MS in normal rat brain tissue and serum were detected. The analysis of variance was applied to compare the intergroup differences of the measurement data. MAIN OUTCOME MEASURES: ① Characteristics of BCNU-PLA-MS release in phosphate buffered solution (PBS) and rat brain tissue; ② Distributions of BCNU-PLA-MS in normal rat brain tissue and serum. RESULTS: ① Release of BCNU-PLA-MS in PBS and rat brain tissue: The BCNU released from BCNU-PLA-MS could be sustained for over 2 weeks both in PBS and brain tissue. In PBS, the released rate of BCNU was over 15% at 24 hours, nearly 50% at 72 hours and over 90% at 15 days. In brain tissue, the released rate was 8% at 4 hours, 16% at 24 hours, 60% at 72 hours, respectively, and BCNU could be sustained released for over 15 days. ② Distributions of BCNU-PLA-MS in normal rat brain tissue and serum: The concentrations of BCNU in the ipsilateral brain tissue were 6 to 70 times higher than those in the contralateral one. The concentrations of BCNU in the ipsilateral brain tissue were obviously higher than those in serum and contralateral brain tissue (F =103.47, P < 0.01). CONCLUSION: BCNU-PLA-MS can increase the drug concentration in targeted brain tissue, decrease that in the non-targeted brain tissue, reduce general toxic and side effects, and have good releasing function.

Spectrofluorimetric Determination of Trace Terbium(III) Using 2,6-Bis-(1'-phenyl-3'-methyl-5'-oxopyrazole-4')pyridinediacyl and N-Cetylpyridium Bromide

A new spectrofluorimetric method for determination of trace terbium based on its reaction with 2.6-his-(1’-phenyl-3’-methyl-5’- pyridinediacyl （H2PMBPP） and N- cetylpyridium bromide （CPB）. at an apparent pH=5.0 provided by a hexamethylenetetramine （5% w/w）-hydrochloric acid buffer,is propesed. The calibration graph is linear in the ran ge

Studies on Semisynthesis and Anibacterial Activity of 7 β-(5-Methyl-1-Aryl-1H-1,2,3-Triazoly-4-Carboxamido) Cephalosporins

Nine new derivatives of 7 β - (5- methyl- 1- aryl- 1H-1, 2, 3- triazoly- 4- carboxamido) cephalosporins were synthesized by acylction of 7 β -amino group of 7-ACA, 7-ADCA and 7-ACT with 5 -methsyl- 1 -aryl- 1 H- 1,2,3-triazoly-4-formyl chloride. The structure of the compounds were confirmed by elementray analysis IR, HNMR and FAB-MS. Some of them showed significant antibacterial

An experimental study on N-ethyl-N-nitrosourea-induced rat brain gliomas

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Synthesis Characterization Non-isothermal Kinetics of the Thermal Decomposition and Redox Properties Derived from Copper(Ⅱ) Binuclear Coordination Compound of 1,4-Bis-(1'-Phenyl-3'-Methyl-5'-Pyrazolone-4')-1,4-Butanedione

The paper reports the synthetic procedure and character of Copper(II) binuclearcoordination compound of 1,4-bis-(1’-phenyl-3’-methyl-5’-pyrazolone Thenon-isothermal kinetics of thermal decomposition of the complex has been stUdied from the TG-DTGcurves by means of the Achar et al. and Coats-Redfern methods,the most probab1e kinetic equation canbe expressed as dofdtrAe -E / RT * l /(2Q).The corresponding kinetic compensation effect expressions arefound to be lnuA=0. 1794E+0. 1689.The non-isothermal thermal decomposition process of the complex isone-dimensional diffusion.But electrochemical studies of the complex(Cu2L’2)from cyclic voltamrnetriccurves by means of powder microelectrodes technique"’,shows one two-electron irreversible process.

亚硝基脲类遗传毒性杂质稳定性影响因素考察

目的:考察以N-甲基-N-亚硝基脲(MNU)和N-乙基-N-亚硝基脲(ENU)为代表化合物的亚硝基脲类遗传毒性杂质的稳定性及其影响因素。方法:采用高效液相色谱(HPLC)考察不同光照(避光、自然光、紫外光)和不同温度(室温、冷藏)条件对ENU和MNU溶液稳定性的影响;采用HPLC考察MNU和ENU在不同储存条件下的短期稳定性;采用气相色谱-质谱联用(GC-MS)考察不同进样口温度(200℃和120℃)对ENU测定准确性的影响。结果:MNU和ENU对光照和温度较为敏感,在光照和室温条件下,均会发生降解;MNU和ENU分别在40℃、75%湿度下放置5天和12天后降解明显;GC-MS进样口温度对ENU的检测影响较大。结论:建议亚硝基脲类遗传毒性杂质应在-20℃避光处保存,在检测过程中控制温度参数;在溶液配制过程中应避光操作、临用新制,以避免出现遗传毒性杂质检测假阴性结果。

ENU诱变构建Jag1基因mRNA剪接异常眼畸形小鼠模型

为建立人类和动物眼病的小鼠模型,采用乙烷基亚硝基脲(ENU)诱导C57BL/6J(B6)小鼠突变手段获得眼病小鼠,利用苏木精-伊红(HE)染色法及免疫组织化学染色技术分析小鼠角膜病变特点,通过连锁分析及位置候选克隆确定致病基因。结果显示,ENU诱变获得1例眼畸形小鼠,小鼠角膜上皮细胞层出现明显空泡,且分化异常,角膜基质层胶原纤维排列杂乱松散,角膜内皮细胞部分脱落。染色体定位将致病基因定位于小鼠第2号染色体微卫星D2Mit107(距着丝粒65.13 cM)与D2Mit423(距着丝粒73.57 cM)之间。对候选基因齿状基因1(Jag1)测序发现,突变杂合子小鼠1条染色体上Jag1基因第24号内含子3′端AG碱基被GG碱基替代,导致第25号外显子存在7个碱基的缺失;蛋白质预测发现,该突变引起Jag1编码区移码及蛋白质编码提前终止。综合分析可知,Jag1基因突变是引起小鼠眼畸形表型的分子基础。

MNU诱导SD大鼠下焦湿热证尿血(膀胱癌)模型研究

目的 探讨N-甲基-N-亚硝基脲(MNU)诱导SD大鼠下焦湿热证尿血——膀胱癌(BC)模型的造模过程和成模率。方法 通过膀胱灌注MNU的方法在造模前、造模中、造模后共计6个时间点进行分批次样品采集,并对膀胱组织进行HE染色以了解膀胱原位癌的形成及发展过程。结果 实验过程顺利,在操作过程中大鼠未出现明显尿路损伤现象。通过本造模手法模型组大鼠膀胱内有明显的上皮增生、破损及大面积肿瘤形成和尿血现象。结论 MNU可以成功诱导SD大鼠下焦湿热证尿血模型(膀胱癌),且成模明显,死亡率低,实验数据可为膀胱癌模型的建立、改良和应用提供一定的参考。

N-甲基-N-亚硝脲诱导的视网膜变性大鼠模型的形态学和功能改变

目的:观察N-甲基-N-亚硝脲(N-m ethyl-N-n itrosourea,MNU)对SD大鼠视网膜光感受器细胞的毒性作用。方法:出生后50 d的SD大鼠84只,随机抽取4只作为正常对照组,余下80只分为4组,每组20只,分别按体质量一次性腹腔注射MNU 80、60、40和30 mg/kg。各组每次4只分别于12 h、24 h、48 h、72 h、120 h行右眼闪光视网膜电图检查(ERG),并于造模后1 d、2 d、3 d、5 d、7 d处死动物,取眼球做组织学检查。结果:1)显示正常对照组的大鼠ERG波形清晰,a、b波振幅正常。MNU 80、60、40和30 mg/kg剂量组a波消失时间分别为3 h、12 h、24 h、120 h;b波消失时间分别为12 h、24 h、72 h、168 h。2)形态学检查测到正常组大鼠中心视网膜的外视网膜厚度为(98.4±1.8)μm。MNU处理后5 d和7 d,80、60、40和30 mg/kg剂量组外视网膜厚度分别为0μm、(9.5±2.3)μm、(42.8±2.7)μm、(71.3±2.4)μm和0μm、0μm、(20.8±2.5)μm、(62.9±2.0)μm,其损伤程度和剂量及时间成正比。结论:MNU可选择性地诱导视网膜光感受器细胞发生凋亡,该作用呈剂量和时间依赖性。

非促分裂haFGF对MNU所致大鼠视网膜损伤的保护作用

目的 研究非促分裂haFGF(nm haFGF)对N 甲基 N 亚硝脲 (MNU)所致大鼠视网膜损伤的保护作用及其作用机制。方法 通过ipMNU(60mg·kg-1 )复制大鼠视网膜损伤模型, 0和 12h后,玻璃体腔内注射不同剂量nm haFGF。24h和 7d后,测量周边视网膜总厚度和外视网膜厚度、光感受器细胞凋亡指数及视网膜Bcl 2和Bax蛋白表达水平。结果 MNU作用 24h后,nm haFGF组周边视网膜的凋亡指数显著降低 (P<0 01); 7d后,nm haFGF组周边视网膜光感受器细胞数明显增多,且周边视网膜总厚度和外视网膜厚度增加 (P<0 01);不同剂量nm haFGF可调节Bcl 2和Bax蛋白表达水平。结论 nm haFGF能部分抑制MNU引起的SD大鼠视网膜损伤,其作用机制可能与上调Bcl 2和下调Bax蛋白表达水平有关。

MNU法诱导大鼠原位膀胱癌模型的建立及MRI的诊断价值

目的:构建N-甲基亚硝基脲(MNU)诱导的大鼠原位膀胱癌模型,探讨利用磁共振成像技术(MRI)对膀胱癌模型进行无创诊断的价值。方法:60只SD雌性大鼠分为实验组45只和对照组15只。实验组大鼠定期膀胱灌注MNU,每次2mg,每2周1次,共4次。对照组大鼠同时膀胱灌注生理盐水,每次0.2mL。于第14周末麻醉大鼠后进行MRI扫描检测,检测后处死大鼠,取膀胱组织进行病理学检测。结果:实验组45只大鼠14周末存活43只,死亡2只,存活率95.6%,死亡率4.4%;经MRI扫描膀胱均见异常信号,提示肿瘤形成,与病理检查结果一致,存活大鼠成瘤率为100%。对照组15只大鼠14周末存活14只,死亡1只,存活率93.3%,死亡率6.7%;经MRI检查未发现膀胱肿瘤,病理检查未见膀胱癌,成瘤率为0。2组大鼠成瘤率比较差异有统计学意义(P<0.05),死亡率比较差异无统计学意义(P>0.05)。结论:MNU膀胱灌注诱导大鼠原位膀胱癌模型方法简便、可靠;MRI扫描结果与病理学检测结果一致,可作为该模型的无创鉴定方法。

不同膳食脂肪酸构成影响MNU诱导大鼠乳腺肿瘤发生的研究

目的:探讨不同膳食脂肪酸对甲基亚硝基脲(MNU)诱导的大鼠乳腺肿瘤发生的影响。方法:用含15%脂肪(wt/wt)的半合成饲料喂养雌性SD大鼠。大鼠50d龄时,分8组喂养8种不同膳食脂肪酸组成的饲料,即SFA、MUFA、n-6PUFA、n-3PUFA、1∶1n-6/n-3、5∶1n-6/n-3、10∶1n-6/n-3和S/M/P为1∶2∶(1其中n-6/n-3为1∶1),每组又分肿瘤组和对喂组,肿瘤组,单次腹腔注射MNU(50mg/kgbw),相应的对喂组注射生理盐水,持续喂养18w。观察各组大鼠体重增长、乳腺肿瘤发生率、肿瘤多发率及肿瘤潜伏期等。结果:除喂养n-3PUFA的大鼠在实验6w后出现体重增长受阻外,其它组别的大鼠体重增长无明显差异。各对喂组大鼠均无乳腺肿瘤发生,而在MNU注射组中,喂养n-3PUFA的大鼠也无乳腺肿瘤发生,但喂养SFA、MUFA、n-6PUFA、5∶1n-6/n-3、10∶1n-6/n-3和S/M/P(1/2/1)大鼠乳腺肿瘤诱发率是1∶1n-6/n-3的2倍,且喂养1∶1n-6/n-3的大鼠,其肿瘤发生的潜伏期也最长。结论:不同膳食脂肪酸构成对MNU诱导的大鼠乳腺肿瘤发生影响不同。,1∶1n-6/n-3膳食脂肪酸构成能有效抑制MNU诱导的乳腺肿瘤发生。