随着经济社会发展，水体富营养化愈发严重，引起蓝藻的异常繁殖而出现水华。蓝藻细胞破裂后能够产生大量的藻毒素，其中的微囊藻毒素(Microcystins, MCs)毒性效应最严重。中华鳖（Pelodiscus sinensis）是广泛分布在我国淡水水体，偏底栖生活的重要经济爬行类物种，MCs进入中华鳖体内后，将对其产生怎样的毒性作用还未见报道。据此，本论文分别探究了急性腹腔注射毒素暴露和慢性养殖水暴露下微囊藻毒素对中华鳖抗氧化防御系统和免疫系统的影响，并初步探讨了中华鳖抗氧化防御和免疫系统之间的联系。

微囊藻毒素暴露实验分为两个。急性毒素暴露实验处理为：将30只中华鳖幼鳖在28±1℃条件下驯化4周，低浓度组和高浓度组分别腹腔注射浓度为0.4 mg/kg体重和2 mg/kg体重的微囊藻毒素（MC-LR），对照组注射等体积生理盐水。处理48 h后取样。慢性毒素暴露实验处理为：30只中华鳖幼鳖驯化4周后，随机分为3组。对照组的养殖水与驯化阶段保持一致，其余两组在MC-LR 20 μg/L的养殖水体中暴露4周，取样对照组和处理组各10只（n=10）。恢复组动物转移到无MC-LR的养殖水体中，继续饲养2周后取样10只。取肝、肾、脾及血液，称重并测量肝、肾、脾、血浆组织中抗氧化防御指标（SOD、CAT、GST活性，GSH、VC、TAOC水平）以及MDA水平；免疫系统指标观测了白细胞分型组成以及血清和肝、脾组织中固有免疫系统C3补体、TNF-α、IL-1β水平和适应性免疫系统IgM含量。

获得主要结果和结论如下：

急性毒性实验中，高浓度组中华鳖肝脏CAT酶活力，肾脏SOD、GST酶活力以及脾脏TAOC水平都显著降低，说明高浓度的MC-LR会降低抗氧化防御功能。而在慢性毒性暴露实验中，处理组和恢复组的肾组织SOD总酶活力均显著低于对照组，这表明MC-LR慢性暴露对肾组织的抗氧化功能产生的影响，经过两周的恢复仍未被消除。

急性毒性实验中，高浓度组肝脏中VC含量以及肾脏中GSH含量显著下降，而肝脏、肾脏和脾脏的MDA含量没有显著改变，说明中华鳖可以耐受高浓度MC-LR，VC和GSH或许在保护中华鳖免受氧化损伤上发挥了重要作用。而在慢性毒性暴露实验中，脾组织中MDA含量显著升高，恢复两周后MDA含量降低至与对照组持平。这说明慢性毒性暴露后，中华鳖脾脏受到氧化损伤，而恢复两周后得到修复。

急性毒性实验中，微囊藻毒素高浓度组的嗜异性粒细胞、嗜酸性粒细胞和淋巴细胞的相对数量发生明显改变；在慢性毒性暴露实验中，嗜酸性粒细胞、淋巴细胞和单核细胞的相对数量变化明显。这说明MC-LR可能是通过影响免疫细胞来干扰中华鳖免疫系统的正常功能。

急性毒性实验中，中华鳖肝脏中TNF-α含量随MC-LR注射浓度的增加呈现明显地先增大后减小的趋势。证明了MC-LR对于中华鳖肝脏固有免疫因子的产生存在低浓度激活、高浓度抑制的二元影响。

在这两种实验处理条件下，中华鳖肝脏指数均显著升高，表明MC-LR会导致中华鳖肝脏肿大。同时，IgM的含量都没有显著改变，表明MC-LR对中华鳖适应性免疫的影响较小。

肝脏和脾脏中大部分抗氧化防御指标和免疫指标之间没有明显的相关性，只有脾脏中MDA和IL-1β呈现统计学显著的弱正相关，TAOC和IL-1β存在统计学显著的弱负相关，表明促炎细胞因子的产生与氧化损伤程度的加深、总抗氧化能力的降低之间有一定程度的联系。

综上，本研究表明，氧化应激参与了微囊藻毒素对中华鳖肝、肾和脾的毒性机制，导致抗氧化防御功能降低，而VC和GSH可以保护中华鳖的这些组织免受氧化损伤。在免疫毒性方面，微囊藻毒素导致肝脏肿大，使白细胞组成发生改变，对固有免疫因子存在二元影响，对适应性免疫反应的影响较小。固有免疫因子与氧化损伤程度以及总抗氧化能力之间存在一定程度的联系。

With the development of society and economy, increasing occurrence of cyanobacterial blooms in eutrophic water bodies has now become a world-wide concern. Cyanobacteria cell can produce phycotoxins, among which microcystins(MCs) are the most  toxic one. Chinese soft-shelled turtle (Pelodiscus sinensis) is an important economic reptile species widely distributed in freshwater bodies in China, which prefers benthic life. The toxicity of MCs on Pelodiscus sinensis is not clear yet. Based on the above background, we explored the effect of microcystins on antioxidant defense system and immune system of Pelodiscus sinensis under acute intraperitoneal injection and chronic microcystins exposure, and preliminarily discussed the relationship between antioxidant defense and immune system of Pelodiscus sinensis in this essay

We designed two microcystins exposure experiments and run respectively. Acute toxicity experiment was managed as: juvenile Chinese soft-shelled turtles 30 individuals were acclimated at 28±1℃ for 4 weeks and divided equally into 3 groups. Turtles in the low and high concentration groups were intraperitoneally injected with 0.4 mg and 2 mg MC-LR·kg^-1 body weight, whereas turtles in the control group were injected with sterile saline. Samples were taken at 48h post-injection. In chronic toxicity experiment, 30 turtles were also randomly divided into three groups. While the control group was raised in water without MC-LR, the other two groups were exposed to water containing MC-LR 20 ug/L for 4 weeks. Then, animals in the recovery group were transferred to water without MC-LR for 2 weeks. Ten turtles were collected from each group (n=10). Liver, spleen, kidney and plasma were taken for measuring morphological indicators and antioxidant defense indexes (activities of SOD, CAT, GST, and content of GSH, VC, T-AOC) and content of MDA. For measuring immune indexes, composition of lymphocytes and content of C3, TNF-α, IL-1β, IgM of the above tissues were determined.

The main results and conclusions are as follows:

1. In acute toxicity experiment, CAT activity of liver, SOD and GST activity of kidney, and TAOC of spleen were significantly decreased in the high concentration group, indicating that high concentration of MC-LR would harm antioxidant defense function. However, in chronic toxicity experiment, SOD activity of kidney in the treatment group and recovery group was significantly lower than that in the control group, indicating that the toxicity of MC-LR chronic exposure on the antioxidant function of kidney did not eliminate after two weeks of recovery.

2. In acute toxicity experiment, VC content in liver and GSH content in kidney decreased significantly in the high concentration group, while MDA content in liver, kidney and spleen did not change significantly, indicating that these tissues did not suffer from oxidative damage. P. sinensis can tolerate high concentrations of MC-LR. VC and GSH may play an important role in protecting from oxidative damage. However, in chronic toxicity experiment, MDA content in spleen increased significantly, and decreased to the control group level two weeks after recovery, indicating that the spleen of Pelodiscus sinensis was damaged by oxidative stress after chronic toxicity exposure, and was repaired after two weeks of recovery.

3. In acute toxicity experiment, the relative numbers of eosinophils, eosinophils and lymphocytes in the high concentration group were significantly changed; The relative numbers of eosinophils, lymphocytes and monocytes changed significantly during chronic toxicity experiment. These results indicated that MC-LR produced obvious immunotoxicity to the leukocyte system, and MC-LR might interfere with the normal function of the immune system by affecting immune cells.

4. In acute toxicity experiment, TNF-α content in the liver showed a trend of first increasing and then decreasing with the increase of MC-LR injection concentration. It was proved that MC-LR had a dualistic effect of low concentration activation and high concentration inhibition on the production of innate immune cytokines.

5. Liver index was significantly increased under the two experimental conditions, indicating that MC-LR could cause enlargement of liver. At the same time, there was no significant change in IgM content, indicating that MC-LR had little effect on adaptive immunity of Pelodiscus sinensis.

6. There was no significant correlation between most of the antioxidant defense indexes and immune indexes in liver and spleen, only MDA and IL-1β had a statistically weak positive correlation; TAOC and IL-1β had a statistically weak negative correlation in spleen. The results indicated that the production of proinflammatory cytokines was related to the severity of oxidative damage and the decrease of TAOC.

In conclusion, oxidative stress was involved in the toxic mechanism of microcystin on liver, kidney and spleen of Pelodiscus sinensis, resulting in decreased antioxidant defense function, while VC and GSH could protect these tissues from oxidative damage. In terms of immunotoxicity, microcystin can cause liver enlargement, change the composition of white blood cells, and have dualistic effects on innate immune cytokines, while have little effect on adaptive immune response. There was a correlation between innate immune factors and the degree of oxidative damage and total antioxidant capacity.