本文選題：microRNA + miR-22　； 參考：《中國農(nóng)業(yè)大學(xué)》2015年博士論文

【摘要】：毛囊是皮膚的衍生物,它的生長發(fā)育具有周期性,即不斷地經(jīng)歷由生長期向退行期和靜止期轉(zhuǎn)變,再由靜止期轉(zhuǎn)向下一個周期的生長期的循環(huán)過程。毛囊從生長期向退行期和靜止期的轉(zhuǎn)變,實際上是一個毛囊退化的生理過程,這個過程受到包括凋亡和生長抑制信號等通路的調(diào)控。但是,生長期末期毛囊細胞分化與毛干裝配形成后毛囊退化過程的具體分子機制還不十分清楚。microRNA(miRNA)是一類20-22個核苷酸大小的非編碼RNA,可以與靶基因的3’UTR結(jié)合而起到抑制靶基因的功能。近年來,越來越多研究報道m(xù)iRNA在毛囊的形態(tài)發(fā)生,毛囊干細胞的增殖、分化和凋亡的穩(wěn)態(tài)平衡等過程中都發(fā)揮著重要作用。已有研究報道表明進化上高度保守的miR-22在退行期和靜止期的表達顯著高于生長期。然而,miR-22是否在毛發(fā)周期和毛囊退化過程中發(fā)揮重要功能需要進一步探討。 本研究首先利用qPCR和原位雜交技術(shù)驗證了miR-22在退行期和靜止期高表達的表達模式。為了進一步揭示miR-22在調(diào)控毛囊發(fā)育和毛發(fā)周期方面的功能,本研究構(gòu)建了可誘導(dǎo)的皮膚特異性過表達miR-22的轉(zhuǎn)基因小鼠K14-rtTA/TRE-miR-22,購買了miR-22敲除鼠。通過體內(nèi)實驗顯示,miR-22過表達可以促進毛囊從生長期向退行期轉(zhuǎn)變,延遲毛囊從靜止期向下一個周期的生長期的轉(zhuǎn)變,從而導(dǎo)致小鼠脫毛。與過表達結(jié)果相反,miR-22敲除導(dǎo)致小鼠毛囊延遲從生長期進入退行期,加快從靜止期向下一個生長期的轉(zhuǎn)變。采用短期誘導(dǎo)miR-22表達的方法證實miR-22可以直接促進毛囊退化過程。進而,揭示miR-22可以通過抑制細胞增殖、細胞遷移、細胞分化和干細胞自我更新,同時促進細胞凋亡的多種途徑來促進毛囊退化過程。 在分子機制方面,本研究通過GO分析發(fā)現(xiàn)miR-22過表達小鼠毛囊生長期(P28,Dox P21)的基因表達譜與野生型小鼠毛囊退行期和靜止期基因表達特征非常相似,即大量毛囊分化相關(guān)的轉(zhuǎn)錄因子基因、角質(zhì)化蛋白基因、非膜結(jié)構(gòu)細胞器基因、凋亡負調(diào)控因子基因以及細胞周期相關(guān)基因都顯著下調(diào)。這說明miR-22是通過下調(diào)這些基因,特別是下調(diào)毛囊分化相關(guān)的轉(zhuǎn)錄因子基因和凋亡負調(diào)控因子基因的表達水平,促進了毛囊退化。為了揭示miR-22如何下調(diào)這些基因,本研究利用生物信息學(xué)方法分析發(fā)現(xiàn)：下調(diào)的毛囊分化相關(guān)的轉(zhuǎn)錄因子和凋亡負調(diào)控因子3'UTR中含有miR-22結(jié)合位點,是miR-22的潛在靶基因。通過雙熒光素酶報告基因檢測實驗進一步驗證了毛囊分化相關(guān)的轉(zhuǎn)錄因子Dlx3、Foxn1和Hoxcl3,以及BMP信號通路抑制因子Sostdc1是miR-22的功能靶基因。這些轉(zhuǎn)錄因子可以直接調(diào)控角蛋白基因的表達來影響內(nèi)根鞘和毛干的形成。綜上所述,本研究揭示miR-22是毛囊退化的重要轉(zhuǎn)錄后調(diào)控因子,可能成為臨床上治療脫發(fā)相關(guān)疾病的靶標(biāo)分子。
[Abstract]:Hair follicles are derivatives of the skin, and their growth and development are cyclical, that is, they are constantly changing from growth phase to receding phase and stationary phase, and then from stationary stage to next cycle. The transition of hair follicles from growth stage to receding stage and stationary stage is actually a physiological process of hair follicle degeneration which is regulated by pathways including apoptosis and growth inhibition signal. However, The molecular mechanism of hair follicle degeneration after the differentiation of hair follicle cells and the formation of hair stem assembly at the end of growth period is not well understood. MicroRNA (miRNA) is a class of non-coding RNAs of 20-22 nucleotides, which can bind to the 3UTR of the target gene and play an inhibitory role. The function of target gene. In recent years, more and more studies have reported that miRNA plays an important role in the process of hair follicle morphogenesis, hair follicle stem cell proliferation, differentiation and homeostasis of apoptosis. It has been reported that the expression of miR-22, which is highly conserved in evolution, is significantly higher in receding and stationary phases than in growth period. However, whether miR-22 plays an important role in hair cycle and hair follicle degeneration needs further study. In this study, qPCR and in situ hybridization techniques were used to verify the high expression patterns of miR-22 in receding and stationary phases. In order to further reveal the function of miR-22 in regulating hair follicle development and hair cycle, a transgenic mouse, K14-rtTA-TRE-miR-22, which can induce skin specific expression of miR-22, was constructed, and miR-22 knockout mice were purchased. In vivo experiments showed that the overexpression of miR-22 could promote the transition of hair follicles from growth stage to receding stage, and delay the transition of hair follicle from stationary stage to next cycle, thus leading to hair loss in mice. Contrary to the overexpression results, the knockout of miR-22 resulted in delayed hair follicle transition from growth stage to retrogression phase, and accelerated the transition from stationary stage to next growth phase. Short-term induction of miR-22 expression showed that miR-22 could directly promote the process of hair follicle degeneration. Furthermore, it is revealed that miR-22 can promote hair follicle degeneration by inhibiting cell proliferation, cell migration, cell differentiation and self-renewal of stem cells, as well as promoting apoptosis. In terms of molecular mechanism, the gene expression profiles of miR-22 overexpression mouse hair follicle growth phase (P28 Dox P21) were similar to those of wild-type mouse hair follicle in receding and stationary stages by go analysis. Many transcription factor genes related to hair follicle differentiation, keratinizing protein genes, non-membrane organelle genes, apoptosis negative regulatory factor genes and cell cycle related genes were significantly down-regulated. This suggests that miR-22 promotes hair follicle degeneration by down-regulating the expression of these genes, especially the transcription factor related to hair follicle differentiation and the expression of apoptosis negative regulator gene. In order to reveal how miR-22 down-regulates these genes, bioinformatics analysis showed that down-regulated transcription factors related to hair follicle differentiation and down-regulated negative regulation factor 3UTR contain miR-22 binding sites, which are potential target genes of miR-22. The detection of double luciferase reporter gene further confirmed that Dlx3 Foxn1 and Hoxcl3, and that BMP signaling pathway inhibitor Sostdc1 is the functional target gene of miR-22. These transcription factors can directly regulate the expression of keratin gene to affect the formation of inner root sheath and hair stem. To sum up, miR-22 is an important posttranscriptional regulator of hair follicle degeneration and may be a target molecule for the treatment of alopecia related diseases.
【學(xué)位授予單位】：中國農(nóng)業(yè)大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2015
【分類號】：R758.7

【參考文獻】

相關(guān)期刊論文 前1條

1 Ganepola AP Ganepola;John R Rutledge;Paritosh Suman;Anusak Yiengpruksawan;David H Chang;;Novel blood-based microRNA biomarker panel for early diagnosis of pancreatic cancer[J];World Journal of Gastrointestinal Oncology;2014年01期

，

本文編號：2063426