Activation of repair pathways after neuronal DNA damage induced by bupivacaine

AIM To investigate the activation of related repair pathways after bupivacaine-induced neuronal DNA damage by cDNA gene screening. METHODS The bupivacaine-induced SH-SY5Y neuronal damage and DNA damage model was established. The technique of cDNA microplate array was used to screen the 21 important regulatory factors in the DNA damage repair pathway. Post-analysis of these differentially expressed repair genes for the repair pathway enrichment and distribution was performed. The data were analyzed by GraphPad Prism 6 statistical software to compare differences between groups. RESULTS The viability of SH-SY5Y cells treated with bupivacaine at different concentrations （detected by CCK-8 assay） showed that the IC₅₀ value of bupivacaine was 1.5 mmol/L. The comet assay related index （the comet tail） was increased （P<0.05）, the phosphorylation level of γH2AX protein was increased （P<0.05）, indicating that DNA damage in the SH-SY5Y cells was significantly aggravated after bupivacaine treatment. The results of cDNA microplate assay showed that compared withcontrol group, the differentially expressed genes after bupivacaine treatment were DNA-PKcs, PTEN, NTH1, RAD9, CSB, GADD45, XPD, XPC-HR23B and P53. The analysis showed that these repair genes were mainly concentrated in the following 3 repair mechanisms： base excision repair, nucleotide excision repair, and non-homologous reconstitution. CONCLUSION The repair genes differentially expressed after neuronal DNA damage caused by local anesthetics are mainly concentrated in the pathways of non-homologous end-joining, base excision repair and nucleotide excision repair.     

苯并[a]芘(BaP)急性暴露对泥蚶消化腺的毒性效应及其应对机制初探

本文研究了苯并[a]芘(benzo[a]pyrene, BaP)急性暴露对泥蚶(Tegillarca granosa)的毒性效应,并以泥蚶的消化腺为目标组织,对泥蚶抵抗BaP毒性的机制进行了初步探究。对样本进行HE染色并对所采集的图像进行分析处理,结果发现,暴露于BaP导致泥蚶消化腺组织出现坏死。对样本进行脂质过氧化并进行DNA氧化损伤测定的结果显示,暴露于BaP导致泥蚶消化腺氧化压力增加,进而导致脂质过氧化以及DNA氧化损伤等细胞层面的损伤。抗氧化酶活性测定结果显示,BaP胁迫诱导泥蚶产生氧化应激反应,抗氧化防御体系积极参与BaP诱导的氧化应激调控过程。神经毒性指标测定结果显示,BaP胁迫可能影响泥蚶神经冲动的传导而具有神经毒性。此外,DNA甲基化水平测定及抗氧化酶基因表达测定的结果显示,泥蚶可能通过降低DNA甲基化水平来激活抗氧化系统以对抗BaP毒性。综上所述,急性BaP暴露会对泥蚶产生显著的毒性效应,以组织损伤、氧化应激反应和神经毒性为主要表征;泥蚶可能通过调整自身DNA甲基化水平来对抗BaP毒性效应。通过本研究,有望为深入探索双壳贝类应对石油污染物胁迫的内在调控机制提供新思路,同时为石油污染威胁下泥蚶的生物资源保护提供参考。     

重金属铬、铜、汞经不同途径神经毒性的对比研究

[目的]探讨几种重金属通过不同的通路对小鼠行为学的影响。[方法]昆明鼠经LashleyⅢH-W水迷宫筛选后,剔除学习记忆差的小鼠,通过慢性滴鼻给药和灌胃的途径,用水迷宫、洞板、自主活动箱测定小鼠给药前后行为学的改变,并利用同步辐射技术,观察小鼠脑部的形态学改变。对通过2种给药途径所造成的神经毒性进行对比研究,探讨重金属通过不同通路对机体所造成的毒性差异及其相关机制。[结果]重铬酸钾滴鼻组用药前后洞板探索行为有明显变化（P〈0.05）,灌胃组给药前后无明显变化;几组重金属滴鼻给药组前后比较,神经系统的兴奋性有所增加,其中重铬酸钾滴鼻组和硫酸铜滴鼻组较为明显,灌胃组给药前后无明显变化;重铬酸钾滴鼻组小鼠和硫酸铜滴鼻组小鼠上台潜伏期有所增长,靶象限活动时间百分比、穿台次数减少,游泳速度有所减慢,灌胃组给药后潜伏期反有所缩短;HE染色和同步辐射观察显示,重铬酸钾滴鼻组小鼠海马部位神经元肿胀明显,空泡变性,可见胶质水肿,部分细胞坏死,并可见呈筛状的坏死灶,胞质中一些区域透光增加,灌胃组无明显改变。[结论]这几种重金属离子在较低浓度时即可通过嗅觉通路沉积于脑,并能通过嗅觉途径改变小鼠的行为学,灌胃组在此浓度时对小鼠的行为学没有较大的改变,有些金属离子可以作为微量元素参与新陈代谢,对神经系统不造成损害。     

Behavioral effects of acute exposure to the insecticide fipronil

The effects of fipronil (Frontline® Top Spot) were investigated in 40 days old rats utilizing open field (OF), hole-board (HB) and elevated plus-maze (EPM) apparatus. Rats (N = 15) received topical application of fipronil (70, 140 and 280 mg/kg) in the neck region and behavior was tested 3 h after administration. Animals treated with corn oil (vehicle) were used as controls. In the OF test animals treated with fipronil at 140 mg/kg showed increased rearing, whereas animals exposed to 280 mg/kg showed increased freezing, grooming, and rearing. In the HB test fipronil at 280 mg/kg increased head-dip and head-dipping behaviors. In the EPM test the only observed effect was increased number of entries in both open and closed EPM arms in animals treated with 280 mg/kg. In conclusion, dermal exposure to fipronil causes effects related to emotionality, fear, and exploratory activity; results add strength to the growing concern that pirazole insecticides can be neurotoxic to humans.     

Behavior changes of learning and memory related to the levels of NO and nNOS in brain of rats with acute alcoholism

AIM: In order to investigate the molecular mechanism of alcoholism acting on learning and memory, the dysfunction of learning and memory function was observed and the content of nitric oxide (NO) and neuronal nitric oxide synthase (nNOS) were determined in rats with acute alcoholism.METHODS: The mature male Sprague-Dawley rats were randomly divided into two groups. The experimental group animals were intraperitoneally administered with ethanol. The control group animals were injected with saline in the same way. The tests of learning and memory were performed at Y-maze after 6 h. Then brains were removed and the content of NO in brain tissue and nNOS expression in hippocampus CA1, corpus striatum were determined, respectively.RESULTS: (1) The training times to reach qualifying standards of Y-maze in experimental group (34.33±13.04) were higher than those in control group (27.50±8.79, P<0.05). (2) The content of NO in experimental group (23.09±9.60) in hippocampus CA1 was significantly increased (P<0.01) compared with that in control group (8.46±5.67). The content of NO in experimental group (19.46±8.25) in corpus striatum was also higher than that in control group (8.22±4.46, P<0.01). (3) The levels of nNOS expression in experimental group (34.33±13.04) in hippocampus CA1 increased significantly (P<0.05) compared with that in control group (27.50±8.79). nNOS positive neurons in experimental group (18.22±7.47) in corpus striatum were also higher than those in control group (10.15±4.24, P<0.05).CONCLUSION: These findings suggest that the mechanism of ethanol neurotoxicity may be partly involved in the signal pathway of NOS and NO in the brain.     

Functional neurotoxic effects in rats acutely exposed to insecticide combinations

The aim of the present study was to analyse the alterations in the cortical and peripheral electrophysiological activity of rats acutely treated with combinations of insecticides. Young adult male Wistar rats were treated with 1/5 and 1/25 LD₅₀ of the insecticides dimethoate, propoxur, cypermethrin and amitraz, given alone or in triple or quadruple combinations. After 24 h, spontaneous cortical activity, and stimulus-evoked cortical and peripheral responses, was recorded and analysed. All treatments changed the cortical activity spectrum. The effect of the 1/5 LD₅₀ combinations indicated non-additive interactions. In the cortical-evoked responses, dimethoate and its combinations gave the strongest change in the latency, while amitraz and its combinations, in the response duration. In the tail nerve, relative refractory period was the most sensitive parameter. The frequency dependence of the cortical responses was the most strongly altered by propoxur, and the least, by amitraz. Our results indicate that simultaneous exposure by various pesticide agents, which happens possibly also in humans, deserves further investigation in, among others, neurotoxicological points of view.