昆明（KM）突变小鼠慢性炎症性皮肤病模型的研究

发布时间：2019-04-10 11:29

【摘要】：背景KM突变小鼠是我所KM封闭群小鼠繁殖过程中出现的一例自发突变小鼠,表型异常,全身被毛稀少,表皮增厚有皱褶。为进一步育种研究,于SPF级动物房内进行近交化培育保种,其突变表型可稳定遗传。该KM突变小鼠初生时与野生型KM小鼠间表型无差异,至一周左右被毛生长后即可与野生型相区分,其被毛稀疏,至离乳仍可见粉红色皮肤,成年后皮肤明显松弛褶皱,表面干燥,部分动物有皮屑脱落,部分动物出现皮肤溃破,类似于人类的慢性炎症性皮肤病病变表现。因此我们对KM突变小鼠进行近交系培育,以期发现其突变的遗传规律并对突变基因进行定位；同时,研究中主要对该突变小鼠的生物学特性进行观察分析,检测其在生物学特性方面与野生小鼠间的差异；根据突变小鼠的表型,观察其皮肤的病理改变及免疫细胞和分子改变,检测免疫器官脾脏和淋巴结淋巴细胞的改变等,目的在于了解KM突变小鼠的系统表型特点、遗传规律等,为其后的基因定位和机制研究奠定基础,以期开发新的疾病动物模型。方法选用同胞杂交、测交等方法近交培育的KM突变小鼠和对照野生小鼠作为研究对象,观察其遗传规律；电子摄像动态观察表型；对不同月龄,不同性别小鼠进行体重、脏器系数、代谢率、体温、及血液学常规、生化检测；通过常规HE病理组织学、免疫组织化学及特殊染色对3月龄、6月龄KM突变小鼠和野生KM小鼠皮肤炎症细胞及细胞因子进行检测比较及脂肪组织脂肪因子瘦素检测比较；对不同年龄段的小鼠皮肤组织、脾脏及淋巴结细胞的凋亡与增殖以及脾脏、淋巴结组织T、B细胞数量进行检测比较。结果KM突变小鼠近交培育至第10代,突变表型稳定遗传,通过观察突变表型外显率,基本符合孟德尔基因分离规律,且与性别无关,推测为单基因控制的常染色体隐性遗传突变；突变表型为皮肤稀毛、皮屑、皮皱等；KM突变小鼠的体重与同龄野生小鼠比较明显降低,且生长缓慢,但除胸腺及腹腔脂肪脏器系数比KM野生小鼠低之外,其它主要脏器重量系数均高于野生KM小鼠；体温高,平均日饮水量大；外周血白细胞、淋巴细胞百分比比同龄野生小鼠多,粒细胞百分比随年龄增长下降；单核细胞百分比呈明显增高的趋势；血糖、总胆固醇、甘油三酯及高密度脂蛋白低、低密度脂蛋白高；皮肤组织病理表现为表皮细胞坏死,上皮角化过度或不全,颗粒层增厚,基底细胞层水肿,真皮浅层血管扩张,结缔组织炎细胞浸润等；皮肤真皮层T细胞、巨噬细胞、肥大细胞浸润,炎症因子IL-6、IL-22、TNF-α、IFN-γ表达增多；脂肪组织瘦素表达增多；表皮细胞凋亡增多,毛囊及皮脂腺细胞增殖减少；脾脏及淋巴结细胞凋亡相对增多,3月龄、6月龄KM突变小鼠增殖增加,9月龄突变小鼠脾脏增殖下降；3月龄、6月龄KM突变小鼠脾脏及淋巴结内T、B淋巴细胞均相对增多,9月龄时脾脏及淋巴结B细胞下降,T细胞增多。结论此次发现的KM自发突变小鼠突变表型主要为皮肤组织自发慢性炎症病变,全身系统性炎症改变,与人类慢性炎症性皮肤病变有相类似的病理改变和细胞分子改变,且表型能稳定遗传。继续深入研究并定位突变基因有望培育成为一种新的慢性炎症性皮肤病的动物模型。
[Abstract]:Background KM mutant mice are one of the spontaneous mutant mice in the breeding process of the KM closed group, and the phenotype is abnormal, the whole body is sparse, and the skin is thickened with wrinkles. In order to further study the breeding, in the SPF animal room, a close-crossing culture is carried out, and the mutant phenotype can be stably inherited. The KM mutant mouse has no difference between the phenotype of the mouse and the wild-type KM mouse, and can be distinguished from the wild-type after a week and so that the KM mutant mouse can be separated from the wild-type after a week, Some of the animals showed skin-breaking, similar to the human chronic inflammatory skin disease. In the same time, the biological characteristics of the mutant mice were observed and analyzed, and the difference between the biological characteristics and the wild mice was detected. according to the phenotype of the mutant mice, the pathological changes of the skin and the changes of the immune cells and the molecules are observed, the changes of the spleen and the lymph node lymphocytes of the immune organs are detected, and the like, Lays a foundation for subsequent gene positioning and mechanism research, with a view to developing new disease animal models. Methods KM mutant mice and control wild mice bred by the method of sibling hybridization and cross-crossing were used as the research object to observe the genetic law of the KM mutant mice and the control wild mice. The dynamic observation of the phenotype was observed. The body weight, the organ coefficient, the metabolic rate, the body weight, the body weight, the organ coefficient, the metabolic rate, the body weight, the body weight, the organ coefficient, the metabolic rate, the body weight, the body weight, the body weight, The results of routine HE, immunohistochemistry and special staining were used to detect the inflammatory cells and cytokines in the skin of three-month-old,6-month-old KM mutant mice and wild KM mice. The apoptosis and proliferation of mouse skin tissue, spleen and lymph node cells in different age groups and the number of T and B cells in the spleen and the lymph node were compared. Results KM mutant mice were bred to the 10th generation and the mutant phenotype was stable, and by observing the apparent rate of the mutant phenotype, it is basically in accordance with the Mendelian gene separation rule, and is not related to sex, and is presumed to be the autosomal recessive genetic mutation of single gene control; and the mutant phenotype is skin. The weight of KM mutant mice was significantly lower than that of the wild mice of the same age, but the weight coefficient of the other major organs was higher than that of the wild KM mice. Large amount of water; the percentage of white blood cells and lymphocytes in the peripheral blood is higher than that of the wild mice of the same age; the percentage of the granulocytes decreases with the age; the percentage of the monocytes is obviously increased; and the blood sugar, the total cholesterol, the triglyceride and the high-density lipoprotein are low, and the low-density lipoprotein high protein; that pathological manifestation of the skin tissue is epidermal cell necrosis, hyperkeratosis or incomplete, thickening of the particle layer, edema of the basal cell layer, superficial vascular expansion of the dermis, infiltration of connective tissue inflammatory cells, etc.; the skin dermal T cell, the macrophage, the mast cell infiltration, the inflammatory factor IL-6, IL, The expression of IL-22, TNF-1, IFN-antigen increased, the expression of leptin in adipose tissue increased, the apoptosis of the epidermal cells increased, the proliferation of the hair follicle and the sebaceous gland decreased, the apoptosis of the spleen and the lymph node was increased, the proliferation of the KM mutant in the 6-month-old and the 9-month-old mutant mice increased, and the spleen of the 9-month-old mutant mice increased. In the 3-month-old and 6-month-old KM mutant, the T and B lymphocytes in the spleen and the lymph nodes of the mice were increased, and the cells of the spleen and the lymph node B were decreased at the age of 9 months, and the T was fine. Conclusion The mutant phenotype of the KM spontaneous mutant mice is mainly the spontaneous and chronic inflammatory disease of the skin, the systemic inflammatory changes of the whole body, the pathological changes and the changes of the cell molecules, and the phenotype of the human chronic inflammatory skin. It can be stably inherited. It is expected that the mutation gene is expected to be cultivated as a new type of chronic inflammatory skin disease
【学位授予单位】：北京协和医学院
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R-332

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