来氟米特治疗子宫内膜异位症大鼠模型的实验研究
发布时间：2018-08-10 20:44
【摘要】：第一部分 非动情期自体移植法子宫内膜异位症大鼠模型的建立 目的：用自体子宫内膜移植法构建子宫内膜异位症大鼠模型,为进一步的实验奠定基础。 方法：健康雌性未孕SD大鼠48只,周龄8-10周,体重200-250g。参考Vernon等的方法进行自体子宫内膜移植。建模后3周进行第二次剖腹探查,观察子宫内膜种植生长情况和组织形态学变化。移植灶体积增大,呈透明结节状或囊状,内有淡黄色透明囊液积聚者视为造模成功。 结果：共做48只SD大鼠模型,有40例大鼠在移植处可见透明小囊肿,表面有血管形成。SD大鼠的内异症模型成模率为83.33%(40/48),有4例大鼠没有成模,考虑在取内膜片段时内膜面积太小或内膜与浆肌层分离不够有关,有2例大鼠死于麻醉意外,还有2例死于腹腔脓肿。其中有10例大鼠异位病灶处腹膜与大网膜粘连,有8例大鼠异位病灶与肠管粘连,余内异症病灶结构清晰,与周围组织无粘连。 结论：非动情期自体子宫内膜移植法建立子宫内膜异位症大鼠模型是可行的。旨在通过这种建模方法建立理想的子宫内膜异位症动物模型,为临床治疗子宫内膜异位症提供实验模型。 第二部分 来氟米特治疗子宫内膜异位症大鼠模型的实验研究 目的：初步探讨来氟米特对EMT大鼠模型异位灶中MAPK及NF-κB的影响,采用酶联免疫法(enzyme-linked immnuno sorbent assay, ELISA)测定大鼠血清中工L-1β的含量,同时观察其对EMT大鼠异位灶体积的抑制作用及对异位内膜病理形态学的影响,并对其可能的机制进行探讨。进一步证实免疫调节剂治疗子宫内膜异位症是可行的,为子宫内膜异位症的临床治疗提供新的治疗策略和靶点以及实验依据。 方法： 1.模型建立3周后,首先抽取实验动物的尾静脉血,用ELISA法检测血清中IL-1β的水平;接着剖腹观察异位子宫内膜的生长情况,测量移植内膜病灶的大小,根据公式体积V=0.52×长×宽×高,计算体积V1,同时把造模成功的40只大鼠随机分为2组：LEF组(n=20)：给予来氟米特35mg/kg/d灌胃,每天同一时间给药,连续用药3周；模型对照组(n=20)：给予生理盐水lml/kg/d灌胃,每天同一时间给药,连续用药3周；给药期间观察大鼠的活动、饮食、排便、及体重变化等情况。 2.喂药3周后,再次抽取实验大鼠的尾静脉血,并剖腹观察大鼠移植内膜病灶的生长情况并测量异位内膜病灶的体积V2,收集移植内膜病灶后处死大鼠：(1)V2与V1进行比较；(2)用免疫组化检测大鼠模型异位灶MAPK及NF-κB的含量；(3)采用ELISA测定大鼠血清中IL-1β的含量；(4)比较各组大鼠治疗前后的体重。 结果： 1.移植模型复制成功,造模成功率83.33%。移植灶体积增大,呈透明结节状或囊状,内有淡黄色透明囊液积聚。 2.用药后LEF组移植内膜病灶体积(54.18±17.67)小于模型对照组(98.03±9.74),差异有统计学意义(P0.05)。 3.用药后LEF组中大鼠移植内膜病灶中MAPK和NF-κB免疫组化平均积分光密度值均较模型对照组低,差异有统计学意义(P0.05)。 4.用药后LEF组血清IL-1β含量明低于模型对照组,差异有统计学意义(P0.05);用药后LEF组血清IL-1β含量明低于用药前,差异有统计学意义(P0.05)。 5.用药后LEF组大鼠行为活跃,毛发光泽度及饮食方面与模型对照组相比均无明显改变。且两组大鼠的体重治疗前后相比,差异无统计学意义(P0.05)。 结论： 1.LEF可使大鼠移植内膜病灶中MAPK和NF-κB的含量降低,使血清IL-1β含量降低。 2.LEF可能通过阻断EMT大鼠模型的MAPK信号转导通路,调控NF-κB通路,影响IL-1β的表达和调控,从而抑制EMT大鼠模型异位子宫内膜的发生及发展,这可能是其治疗EMT的机制之一。
[Abstract]:Part one
Establishment of a rat model of endometriosis induced by non estrous autotransplantation
Objective: To establish a rat model of endometriosis by autologous endometrial transplantation and lay a foundation for further experiment.
Methods: 48 healthy female sterile SD rats, aged 8-10 weeks and weighing 200-250g, underwent autologous endometrial transplantation with reference to Vernon et al. The second exploratory laparotomy was carried out 3 weeks after the establishment of the model. The growth and histomorphological changes of endometrial implantation were observed. Cystic fluid accumulation is regarded as a successful model.
Results: A total of 48 SD rat models were made, 40 of which showed small transparent cysts and angiogenesis on the surface of the graft. The modeling rate of endometriosis in SD rats was 83.33% (40/48), and 4 rats were not. Considering that the intimal area was too small or the intimal membrane was not separated from the serosomal layer, 2 rats died of anesthesia accident. Among them, 10 rats had adhesion between peritoneum and omentum, 8 rats had adhesion between ectopic lesion and intestine, and the remaining endometriosis had clear structure and no adhesion with surrounding tissues.
CONCLUSION: It is feasible to establish a rat model of endometriosis by autologous endometrial transplantation in the non-estrous period.
The second part
Experimental study of leflunomide in the treatment of endometriosis in rats
AIM: To investigate the effects of leflunomide on MAPK and NF-kappa B in ectopic foci of EMT rats, and to determine the content of working L-1 beta in serum by enzyme-linked immnuno sorbent assay (ELISA), and to observe the inhibition of leflunomide on the volume of ectopic foci and the pathomorphological changes of ectopic endometrium in EMT rats. The possible mechanism is discussed. It is further proved that immunomodulators are feasible in the treatment of endometriosis and provide new therapeutic strategies, targets and experimental basis for the clinical treatment of endometriosis.
Method:
1. Three weeks after the establishment of the model, the tail vein blood of the experimental animals was firstly extracted and the level of IL-1 beta in the serum was detected by ELISA. Then the growth of ectopic endometrium was observed by laparotomy and the size of the transplanted endometrial lesion was measured. According to the formula volume V = 0.52 * length * width * height, the volume V1 was calculated. At the same time, 40 successful rats were randomly divided into two groups. LEF group (n=20): Leflunomide 35 mg/kg/d was given intragastric administration at the same time every day for 3 weeks; model control group (n=20): normal saline lml/kg/d was given intragastric administration at the same time every day for 3 weeks; rats'activities, diet, defecation and weight changes were observed during the administration.
2. After 3 weeks, the tail vein blood of experimental rats was extracted again, and the growth of endometrial lesions was observed by laparotomy, and the volume of ectopic endometrial lesions was measured by V2. The rats were sacrificed after endometrial lesions were collected: (1) V2 and V1 were compared; (2) The contents of MAPK and NF-kappa B in the ectopic lesions of the rat model were detected by immunohistochemistry; (3) ELIS was used to detect the volume of ectopic endometrial lesions. A was used to determine the content of IL-1 beta in rat serum; (4) the weight of rats in each group was compared before and after treatment.
Result:
1. The transplantation model was successfully reproduced with a success rate of 83.33%. The volume of the graft was enlarged, with clear nodular or cystic shape and light yellow cystic fluid accumulation.
2. The volume of transplanted endometrial lesions in LEF group was smaller than that in model control group (54.18
3. The mean integral optical density of MAPK and NF-kappa B in LEF group was lower than that in model control group (P 0.05).
4. The level of serum IL-1beta in LEF group was significantly lower than that in model control group (P 0.05), and the level of serum IL-1beta in LEF group was significantly lower than that before treatment (P 0.05).
5. After treatment, the behavior of LEF rats was active, and there were no significant changes in hair gloss and diet compared with the model control group. There was no significant difference in body weight between the two groups before and after treatment (P 0.05).
Conclusion:
1. LEF can decrease the contents of MAPK and NF-kappa B in endometrial lesions of rats, and decrease the content of IL-1 beta in serum.
2. LEF may inhibit the development of ectopic endometrium in EMT rats by blocking MAPK signal transduction pathway, regulating NF-kappa B pathway, affecting the expression and regulation of IL-1 beta, which may be one of the mechanisms of LEF in treating EMT.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R-332;R711.71
本文编号：2176152
[Abstract]:Part one
Establishment of a rat model of endometriosis induced by non estrous autotransplantation
Objective: To establish a rat model of endometriosis by autologous endometrial transplantation and lay a foundation for further experiment.
Methods: 48 healthy female sterile SD rats, aged 8-10 weeks and weighing 200-250g, underwent autologous endometrial transplantation with reference to Vernon et al. The second exploratory laparotomy was carried out 3 weeks after the establishment of the model. The growth and histomorphological changes of endometrial implantation were observed. Cystic fluid accumulation is regarded as a successful model.
Results: A total of 48 SD rat models were made, 40 of which showed small transparent cysts and angiogenesis on the surface of the graft. The modeling rate of endometriosis in SD rats was 83.33% (40/48), and 4 rats were not. Considering that the intimal area was too small or the intimal membrane was not separated from the serosomal layer, 2 rats died of anesthesia accident. Among them, 10 rats had adhesion between peritoneum and omentum, 8 rats had adhesion between ectopic lesion and intestine, and the remaining endometriosis had clear structure and no adhesion with surrounding tissues.
CONCLUSION: It is feasible to establish a rat model of endometriosis by autologous endometrial transplantation in the non-estrous period.
The second part
Experimental study of leflunomide in the treatment of endometriosis in rats
AIM: To investigate the effects of leflunomide on MAPK and NF-kappa B in ectopic foci of EMT rats, and to determine the content of working L-1 beta in serum by enzyme-linked immnuno sorbent assay (ELISA), and to observe the inhibition of leflunomide on the volume of ectopic foci and the pathomorphological changes of ectopic endometrium in EMT rats. The possible mechanism is discussed. It is further proved that immunomodulators are feasible in the treatment of endometriosis and provide new therapeutic strategies, targets and experimental basis for the clinical treatment of endometriosis.
Method:
1. Three weeks after the establishment of the model, the tail vein blood of the experimental animals was firstly extracted and the level of IL-1 beta in the serum was detected by ELISA. Then the growth of ectopic endometrium was observed by laparotomy and the size of the transplanted endometrial lesion was measured. According to the formula volume V = 0.52 * length * width * height, the volume V1 was calculated. At the same time, 40 successful rats were randomly divided into two groups. LEF group (n=20): Leflunomide 35 mg/kg/d was given intragastric administration at the same time every day for 3 weeks; model control group (n=20): normal saline lml/kg/d was given intragastric administration at the same time every day for 3 weeks; rats'activities, diet, defecation and weight changes were observed during the administration.
2. After 3 weeks, the tail vein blood of experimental rats was extracted again, and the growth of endometrial lesions was observed by laparotomy, and the volume of ectopic endometrial lesions was measured by V2. The rats were sacrificed after endometrial lesions were collected: (1) V2 and V1 were compared; (2) The contents of MAPK and NF-kappa B in the ectopic lesions of the rat model were detected by immunohistochemistry; (3) ELIS was used to detect the volume of ectopic endometrial lesions. A was used to determine the content of IL-1 beta in rat serum; (4) the weight of rats in each group was compared before and after treatment.
Result:
1. The transplantation model was successfully reproduced with a success rate of 83.33%. The volume of the graft was enlarged, with clear nodular or cystic shape and light yellow cystic fluid accumulation.
2. The volume of transplanted endometrial lesions in LEF group was smaller than that in model control group (54.18
3. The mean integral optical density of MAPK and NF-kappa B in LEF group was lower than that in model control group (P 0.05).
4. The level of serum IL-1beta in LEF group was significantly lower than that in model control group (P 0.05), and the level of serum IL-1beta in LEF group was significantly lower than that before treatment (P 0.05).
5. After treatment, the behavior of LEF rats was active, and there were no significant changes in hair gloss and diet compared with the model control group. There was no significant difference in body weight between the two groups before and after treatment (P 0.05).
Conclusion:
1. LEF can decrease the contents of MAPK and NF-kappa B in endometrial lesions of rats, and decrease the content of IL-1 beta in serum.
2. LEF may inhibit the development of ectopic endometrium in EMT rats by blocking MAPK signal transduction pathway, regulating NF-kappa B pathway, affecting the expression and regulation of IL-1 beta, which may be one of the mechanisms of LEF in treating EMT.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R-332;R711.71
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