大鼠胚胎卵巢冷冻保存和异体无血管皮下移植的生长

背景：胚胎卵巢组织免疫原性低,不易引起宿主的免疫排斥反应,是卵巢异体移植的理想供体。但由于获取胎儿卵巢的不随意性和来源限制,标本的冷冻显得至关重要,成功的冷冻保存和复苏技术是建立胎儿卵巢库的关键。 目的：通过对大鼠新鲜或冻融胚胎卵巢组织异体无血管移植后的组织形态和功能的观察,探讨胚胎卵巢组织异体移植和超速冷冻保存的可行性。 设计、时间及地点：随机对照动物实验,于2006-03/05在云南省第一人民医院生殖遗传科进行课题设计,于2006-05/2007-02在云南大学单克隆抗体研究中心完成实验。 材料：卵巢来源于孕龄为18~20 d SD大鼠胚胎。取阴道脱落细胞涂片证实有正常动情周期未交配过的SD成年雌性大鼠作为移植对象,分4组：正常对照组（n=10）、去势对照组（n=10）、新鲜胚胎卵巢移植组（n=25）、冷冻胚胎卵巢移植组（n=25）。 方法：正常对照组仅行皮肤筋膜肌肉切开缝合术,去势对照组切除双侧卵巢组织。将孕龄为18~20 d的新鲜或经超速冷冻法冷冻复苏的大鼠胚胎卵巢植入去势后4周的大鼠颈部皮下作为实验组,分别为新鲜移植组和冷冻移植组。 主要观察指标：通过阴道脱落细胞的观察和血清雌二醇、孕酮的测定了解大鼠内分泌恢复情况,并对移植卵巢进行组织学观察。 结果：新鲜移植组和冷冻移植组分别有52.17%和38.01%大鼠恢复了动情周期,两组血清雌二醇和孕酮在分别在35和49 d恢复到正常,且动情周期天数与正常对照组比较,差异无显著性意义。冷冻移植组大鼠恢复动情周期虽稍短于新鲜移植组,但差异并无统计学意义。63 d后取出移植物观察,见移植物色泽红润,表面有丰富的血管形成,镜下见卵巢内有大量的小动脉和小静脉;移植物组织学检查发现卵泡的发育,光镜下可见不同发育阶段的卵泡,有成熟卵泡和黄体形成,形态与正常卵巢无明显差别。 结论：大鼠胚胎卵巢异体异位无血管皮下移植后能存活、生长发育并具有与正常卵巢相似的内分泌功能;超速冷冻法能较好的保存胚胎卵巢的活性,复苏后与新鲜胚胎卵巢组织具有相同的生理活性。     

长期力竭运动动情周期抑制模型大鼠卵巢组织形态和超微结构的变化

目的: 观察运动致大鼠卵巢的组织形态和细胞超微结构的变化。 方法: 将正常的20只SD雌性大鼠（3月龄）随机分为对照组和造模组（各10只），其中造模组大鼠每日进行一次力竭游泳运动, 直至其动情周期紊乱，然后观察其卵巢组织形态和细胞超微结构的变化。 结果：与对照组相比，造模组大鼠卵巢典型生长卵泡和新鲜黄体明显减少，原始卵泡、闭锁卵泡则相对增加；其颗粒细胞、卵泡膜细胞、黄体细胞内细胞器明显减少而含有大量的脂滴。 结论: 长期力竭运动可抑制大鼠卵巢卵泡的发育、成熟、排卵与黄体形成，并使卵巢细胞的超微结构发生抑制性改变。     

仙珍骨宝胶囊对去卵巢大鼠股骨颈骨代谢的影响*☆◆

背景：用骨组织形态计量学方法探讨中药对去卵巢大鼠股骨颈骨质疏松的影响,可为采用中药防治绝经后妇女骨质疏松性股骨颈骨折提供实验依据。 目的：观察仙珍骨宝胶囊对去卵巢大鼠股骨颈松质骨的影响。 方法：3月龄SD雌鼠随机分为4组：基础对照组于实验开始时处死取材,去卵巢组和仙珍骨宝组去卵巢造模,仙珍骨宝组在去卵巢后灌胃仙珍骨宝,去卵巢组和年龄对照组灌胃生理盐水,90 d后处死,取股骨颈经不脱钙骨制片进行骨组织形态计量学参数测量。 结果与结论：与年龄对照组比较,去卵巢大鼠静态参数的骨小梁面积百分数和骨小梁数量明显减少(P < 0.01),骨小梁间隙明显增大(P < 0.01);动态参数的每毫米破骨细胞数和破骨细胞贴壁表面长度明显增加(P < 0.01),骨矿化沉积率明显减少(P < 0.01)。说明去卵巢能导致大鼠股骨颈骨量显著减少。给予仙珍骨宝治疗后,大鼠的骨小梁厚度及骨小梁面积明显增加(P < 0.05),每毫米破骨细胞数和破骨细胞贴壁表面长度有所减少,标记周长百分数则有所增加。说明仙珍骨宝能阻止去卵巢所致的大鼠股骨颈骨量丢失。     

海藻酸钠-多聚赖氨酸-海藻酸钠微囊化大鼠卵巢细胞腹腔异体移植以及

背景：应用海藻酸钠-多聚赖氨酸-海藻酸钠微囊技术包裹细胞进行异体移植,被证明是一种可避免受体产生免疫排异的有效方法。 目的：观察海藻酸钠-多聚赖氨酸-海藻酸钠微囊化卵巢细胞种植于去卵巢大鼠腹腔后对受体大鼠肾上腺的影响。 设计、时间及地点：随机对照动物实验,于2007-03/2008-09在首都医科大学生殖医学研究中心完成。 材料：取Wistar大鼠卵巢分离培养卵巢细胞,进行微囊化处理,制备包裹卵巢细胞的海藻酸钠-多聚赖氨酸-海藻酸钠微囊。 方法：将40只雌性Wistar大鼠随机分为4组：去势组于无菌条件下切除双侧卵巢;微囊移植组手术摘除双侧卵巢后腹腔移植包裹卵巢细胞的海藻酸钠-多聚赖氨酸-海藻酸钠微囊;雌激素替代治疗组手术摘除双侧卵巢后腹腔注射乙烯雌酚 0.2 mg/kg,每3 d注射1次。正常对照组不进行任何处理。 主要观察指标：干预后30 d放射免疫法检测大鼠血清中雌激素水平,观察肾上腺切片的形态学特点,并利用免疫组织化学SP法检测各组大鼠肾上腺皮质各层增殖细胞核抗原表达变化。 结果：①放射免疫法检测正常对照组,雌激素替代治疗组以及微囊移植组雌激素水平明显高于去势组,而正常对照组,微囊移植组,雌激素替代治疗组之间雌激素水平差异无显著性意义。②去势组肾上腺皮质网状带增厚,网状带与球束状带比值增大,束状带和网状带增殖细胞核抗原阳性表达都增加;而微囊移植组及雌激素替代治疗组增殖细胞核抗原阳性细胞明显低于去势组,差异具有显著性意义。 结论：微囊化大鼠卵巢细胞异体移植后,可在受体大鼠体内继续合成和分泌雌二醇,所分泌的雌激素可以纠正肾上腺皮质因卵巢摘除而导致的形态改变,并与雌激素替代治疗疗效相近。     

癫对雌性大鼠生殖内分泌的影响

目的探讨癫对雌性大鼠生殖内分泌的影响。方法将动情周期规律的成年雌性Wistar大鼠30只随机分为对照组、模型组,每组各15只。模型组大鼠腹腔注射氯化锂-匹罗卡品建立癫模型,对照组大鼠腹腔注射生理盐水,通过阴道脱落细胞涂片判定两组大鼠动情周期的变化,用在四周后在动情间期处死、采血,采用放免法测定血清性激素水平,分离子宫、卵巢组织,检测大鼠体质量、子宫、卵巢重量及子宫、卵巢组织学结构。结果对照组15只大鼠无癫发作,无死亡,动情周期稳定,模型组有3只死亡,其余12只大鼠造模成功,表现为动情周期紊乱,血清性激素中黄体生成素及孕酮水平下降,体质量减轻,子宫和卵巢重量下降,光镜检查子宫腺体数量变少,卵泡变少,与对照组比较差异均有统计学意义（P均〈0.05）。结论癫对雌性Wistar大鼠生殖发育有损害作用。     

中国雪蛤酶解液预防去卵巢大鼠骨质疏松

背景：以往研究发现雪蛤具有预防绝经后相关疾病的作用,但是否具有预防绝经后骨质疏松的功效有待于深入研究。 目的：验证雪蛤酶解液对去卵巢大鼠骨质疏松的预防作用。 方法：健康成年Wistar大鼠72只,按随机数字表法分为6组,每组12只。假手术组大鼠不切除卵巢仅切除卵巢周围部分脂肪,其余各组均切除卵巢造成骨质疏松模型。于造模后第7天开始,各组等容灌胃,连续12周。假手术组和模型组每日灌胃等量蒸馏水;普雷马林组每日灌胃387.8 g/L普雷马林(结合雌激素片)悬浮液6. 67 g/(kg•d);67.0,26.8,13.4 g/(kg•d) 蛤士蟆酶解液组每日分别按相应剂量灌胃。每个月测定食物利用率;实验12周末时,处死大鼠后取子宫,测定子宫质量;取左、右侧股骨和腰椎骨,双能X射线骨密度仪测定骨密度;电感偶合等离子体直读光谱仪测定骨钙、磷含量;脱脂、灰化后测定骨干质量、灰质量。 结果与结论：在4组干预组中,67.0 g/(kg•d)蛤士蟆酶解液组和普雷马林组的食物利用率最低(P < 0.05);在各切卵巢组中,普雷马林组的子宫质量最重(P < 0.05);26.8 g/(kg•d) 蛤士蟆酶解液和普雷马林可以延缓切卵巢大鼠股骨骨密度的下降(P < 0.05);67.0,26.8,13.4 g/(kg•d)蛤士蟆酶解液组骨钙和骨磷与模型组比较均有明显提升(P < 0.05),与普雷马林组差异无显著性意义(P > 0.05)。提示雪蛤酶解液对去卵巢大鼠的骨质疏松具有预防作用,其疗效与结合雌激素片相当,且对子宫增生的不良反应低于结合雌激素片。     

去卵巢牙周炎模型大鼠牙槽骨吸收与补骨脂的干预

背景：补骨脂在治疗骨质疏松症方面有明显效果。 目的：观察补骨脂对去卵巢牙周炎大鼠牙槽骨代谢、牙槽骨密度及高度的影响。 方法：将30只雌性Wistar大鼠随机等分为3组：去卵巢组和补骨脂组行双侧卵巢切除术,假手术组仅手术不切除卵巢,补骨脂组于卵巢切除第2天灌胃3 g/kg补骨脂水剂,1次/d。去卵巢2周,所有大鼠采用结扎上颌磨牙的方法建立牙周炎模型。 结果与结论：上颌磨牙结扎12周,与假手术组比较,去卵巢组大鼠血清雌二醇、钙离子浓度显著降低(P < 0.05),碱性磷酸酶水平显著升高(P < 0.05),X射线片及苏木精-伊红染色显示去卵巢组大鼠上颌骨骨密度及高度显著降低(P < 0.05);与去卵巢组比较,补骨脂组大鼠血清钙离子浓度显著升高(P < 0.05),碱性磷酸酶水平显著降低(P < 0.05),上颌骨骨密度及高度显著增高(P < 0.05),且与假手术组比较差异无显著性意义(P > 0.05)。说明雌激素缺乏促进实验性牙周炎大鼠牙槽骨吸收,补骨脂可有效阻止雌激素降低导致的牙槽骨高度、骨密度降低。     

螺内酯对碱烧伤诱导大鼠角膜新生血管形成的影响及其机制研究

背景：螺内酯为醛固酮受体拮抗剂,近来有实验证实螺内酯在体内外能够有效地抑制新生血管的形成。 目的：验证螺内酯对碱烧伤诱导大鼠角膜新生血管的抑制作用。 方法：取SD大鼠36只,采用碱烧伤方法制备大鼠角膜新生血管模型,造模后以数字表法随机分成实验组和对照组。实验组术后灌胃给予螺内酯100 mg/kg,1次/d,对照组灌胃给予等量生理盐水。另取大鼠6只不作任何处理作为正常对照组。于造模后4,7,14 d运用裂隙灯观察各组大鼠角膜新生血管并计算面积,并每组随机处死6只大鼠,运用免疫组化染色方法和计算机图像分析系统检测观察血管内皮细胞生长因子及基质金属蛋白酶2的表达。 结果与结论：正常角膜组织未见炎症细胞与新生血管,角膜上皮及基质层仅见微弱血管内皮细胞生长因子表达,未见基质金属蛋白酶2表达。与对照组比较,实验组各个时期新生血管面积减小,血管内皮细胞生长因子和基质金属蛋白酶2表达降低(P < 0.05)。提示螺内酯可能通过参与下调血管内皮生长因子和基质金属蛋白酶2表达,从而有效地抑制角膜新生血管的形成。     

雌激素对去卵巢大鼠缺血再灌注脑损伤的保护作用

目的探讨雌激素对去卵巢大鼠缺血再灌注脑损伤的保护作用。方法将大鼠切除双侧卵巢后30d给予肌肉注射苯甲酸雌二醇100μg/(kg·d)连续14d,然后制作大鼠局灶性脑缺血再灌注模型;制模12h后取脑组织行免疫组化染色,检测细胞间黏附分子(CD54)、肿瘤坏死因子α(TNF-α)的表达;TUNEL法检测脑组织凋亡细胞数;电镜观察脑细胞膜超微结构的变化。结果与缺血再灌注组及去卵巢组比较,雌激素组脑组织CD54、TNF-α表达明显降低,凋亡细胞数明显减少(均P<0.05);脑细胞膜结构非特异性损伤减轻。结论雌激素通过减少缺血再灌注大鼠脑组织炎性细胞因子表达、减轻炎症反应、降低脑组织细胞凋亡而发挥脑保护作用。     

雌激素与辛伐他汀序贯法干预对去势大鼠骨质疏松的治疗作用

背景：Frost根据骨重建的概念创建了骨重建干预理论——序贯疗法,即在骨吸收抑制剂之后可给予刺激骨形成的药物。 目的：基于骨重建干预理论,序贯应用雌激素与辛伐他汀干预骨重建吸收期和形成期,观察其对去势大鼠骨质疏松的治疗作用。 方法：3月龄雄性SD大鼠40只,以随机数字表法分为去势组与正常对照组。去势组切除双侧卵巢,正常对照组只进行下腹部皮肤单纯切开术。大鼠去势后1个月,将去势组随机分为3组：序贯组、雌激素组、去势对照组,并开始药物干预：序贯组,皮下注射苯甲酸雌二醇0.1 mg/kg,3 d给药1次,2周后,灌胃给予辛伐他汀5 mg/(kg•d)2周,停药5周,再应用辛伐他汀5 mg/(kg•d)灌胃2周;雌激素组,皮下注射苯甲酸雌二醇0.1 mg/kg,每3 d给药1次,连续用药11周;去势对照组,单纯的饲料喂养,无药物干预。11周后,双能X射线骨密度仪测定股骨骨密度,放射免疫法检测血清白细胞介素6、骨钙素水平。 结果与结论：各治疗组大鼠股骨骨密度、骨钙素水平高于去势对照组(P < 0.05),并且序贯组明显高于雌激素组(P < 0.05)。各治疗组白细胞介素6水平低于去势对照组(P < 0.05),并且序贯组低于雌激素组(P < 0.05)。说明雌激素和辛伐他汀序贯疗法可以通过抑制骨吸收,促进骨形成有效地治疗骨质疏松。     

Neural modulation of ovarian function

To many neuroendpcrinologists, all that an animal does to bring about ovulation is to produce a surge of luteinizing hormone, which, in its turn, acts on an ovary containing ripe follicles. In recent years, the evidence has been strengthening in favour of a direct nervous input to the ovary which, at the very least, modulates the humoral hormonal stimulus for ovulation. Some of this evidence is presented below.     

Neural participation in ovarian control

In this article, Ida Gerendai and Béla Halász describe the elegant studies from which they have derived the evidence for the existence and pathways of efferent and afferent neural connections between the ovary and hypothalamus, and the role which these pathways have to play in gonadal-hypothalamic feedback. The model which they have used concerns the effects of compensatory ovarian hypertrophy following hemiovariectomy. For the neuroendocrinologist, the involvement of possible neural pathways in areas previously considered to be only under a humoral control, has many far-reaching implications, and may provide explanations for a number of previously inexplicable observations.     

Neural participation in ovarian control

In this article, Ida Gerendai and Béla Halász describe the elegant studies frr which they have derived the evidence for the existence and pathways of efferent and afferent neural connections between the ovary and hypothalamus, and the role which these pathways have to play in gonadal-hypothalamic feedback. The model which they have used concerns the effects of compensatory ovarian hypertrophy following hemiovariectomy. For the neuroendocrinologist, the involvement of possible neural pathways in areas previously considered to be only under a humoral control, has many far-reaching implications, and may provide explanations for a number of previously inexplicable observations.     

Effect of secretion of splenocytes after superior ovarian nerve section on the ovarian steroidogenesis.

It is known that ovary and spleen are innervated extensively by afferent and efferent noradrenergic sympathetic nerve fibers from the celiac ganglion. Furthermore, immune cells located in the ovary influence the ovarian physiology. However, the peripheral interaction between the immune and neuroendocrine system is poorly understood. This work was undertaken to study the effect of superior ovarian nerve (SON) transection, in adult rats, on the number of splenocyte beta-adrenergic receptors and their possible relation to ovarian steroidogenesis, measuring the effect of secretions of those splenocytes on progesterone and estradiol release from the ovary. Seven days after SON transection, the splenocytes were isolated and then cultured for 48 h. Their number of beta-adrenergic receptors, measured using [125I]-cyanopindolol as ligand, increased, and their culture media, used to stimulate ovaries from 60-day-old intact (neither SON-transected nor sham-operated) rats in vitro on diestrous day 2 showed a decrease in progesterone release and an increase in estradiol release in relation to splenocyte culture media of control rats (sham-operated; p < 0.001, respectively). The effects of in vivo SON transection were simulated by an in vitro system modulating the splenocyte beta-adrenergic receptor number. The splenocytes from SON-transectioned rats were preincubated with and without norepinephrine (NE) 10(-6) M for 48 h, a low and high number of beta-adrenergic receptors respectively, and then were stimulated with NE 10(-6) M for 24 h. After that, the culture medium from splenocytes with a low number of beta-adrenergic receptors induced progesterone release from the ovaries of intact rats (p < 0. 001), but produced no change in estradiol release. The data suggest that splenocyte secretions, which participate in the ovarian steroidogenic response, particularly in progesterone release, might be controlled by adrenergic influences since the number of splenocyte beta-adrenergic receptors changes through SON-celiac ganglion-noradrenergic postganglionic innervation of the spleen. In estradiol release, probably other neurotransmitters than norepinephrine (NE) are involved when the SON is sectioned. In this paper we also show functional evidence for modulation of immune function by the sympathetic nervous system and its principal neurotransmitter, NE.     

Paraneoplastic cerebellar degeneration associated with ovarian cancer: anti-Yo immunoreactivity in autoptic cerebellum and ovarian carcinoma

Paraneoplastic cerebellar degeneration is a rare disorder caused likely by autoimmune mechanisms in malignant oncologic diseases, and the most common tumors are ovarian, breast, lung cancer, and m. Hodgkin. An immune reaction is supposed to be directed against identical antigens of cerebellum and tumor, and paraneoplastic antibodies called anti-Yo, anti-Hu, anti-Ri, or anti-Tr are often detected in blood and cerebrospinal fluid. The course of paraneoplastic cerebellar degeneration as a complication of ovarian cancer is described. The relationship between the malignancy and pathologic changes in cerebellum was confirmed by positive immunohistochemical and immunofluorescence reaction between a patient's anti-Yo-positive serum and her own Purkinje's and ovarian cancer cells.     

Myasthenia gravis and premature ovarian failure

We describe a patient who developed seropositive myasthenia gravis 16 years after she was diagnosed with autoimmune premature ovarian failure with antibodies to the receptor for follicle-stimulating hormone (FSH). Although thymectomy led to improvement of her myasthenic symptoms, menses did not resume. Such combined seropositivity for antibodies to acetylcholine and ovarian hormone receptors in a patient with myasthenia gravis and premature ovarian failure may reflect common disease mechanisms, although the precise pathogenesis of these disorders remains ill-defined.     

Leukodystrophy in patients with ovarian dysgenesis

We describe clinical, biochemical, pathological, and spectroscopic findings in 4 women, aged 15 to 29 years, from three unrelated families who had a unique combination of a central neurvous system white matter disease and primary ovarian failure. All had normal initial development but 3 had borderline low IQ and academic difficulties in primary school. Puberty did not develop in 2 patients and was arrested in a third patient. The fourth patient had premature ovarian failure at the age of 13 years. Head magnetic resonance imaging showed diffuse white matter disease, with frontal cortical atrophy in the most clinically advanced patient. All patients had normal karyotype and normal findings on extensive evaluations for known leukodystrophies, for other metabolic diseases, and for causes of ovarian failure. Proton magnetic resonance spectroscopic imaging showed reduction of choline-containing compounds in the affected white matter in all patients and reduction of N-acetylaspartate in the unaffected frontal white matter of 2 patients. All patients had evidence of primary gonadal insufficiency with a normal hypothalamic-hypophyseal axis. Pathological analysis showed streak ovaries in 1 patient and signs of hypomyelination, and gliosis on brain biopsy in another patient. In conclusion, we present a novel group of patients who have in common leukodystrophy, primary ovarian dysfunction, and magnetic resonance spectroscopic abnormalities.