小分子核糖核酸-137對食慾及代謝之影響造成小鼠體重增重

Abstract

小分子核醣核酸(Micro RNA, miRNA)是一段長度約22個核苷酸的RNA分子，目前已被證實會透過和目標基因的結合影響其表達。本次研究所探討的miR-137，除了已知在腦部具有很高的表現量，並會參與神經元的分化與成熟，已經有團隊透過全基因組關聯分析(Genome-wide association analysis, GWAS)發現和神經官能症(Schizophrenia)相關的單核苷酸多型性(Single-nucleotide polymorphism, SNP)就位於miR-137的上游，另外也有許多文獻探討miR-137在癌症所扮演的角色，像是在非小細胞肺癌(Non-small cell lung cancer, NSCLC)中被認為可以用來預測病人具有不好的預後。miR-137廣泛地參與許多生物體的作用，於是我們以此為基礎，想要進一步在動物模型探討miR-137的生理意義。首先我們針對小鼠的各個重要器官進行miR-137表現量之剖析，發現前三高表現量的器官分別是大腦、心臟及肝臟，考量實驗室所擁有的方法學，我們選擇阿黴素（Doxorubicin）、乙醯胺酚（Acetaminophen）、脂多醣（Lipopolysaccharide）、高油脂飼料（High-fat diet）分別針對心臟及肝臟（後三者）給予小鼠短期壓力，結果發現只有在經過一個月的高油脂飼料的餵食後，小鼠肝臟的miR-137表現量顯著地下降了。而在先前實驗室所製作出的miR-137基因全身剔除小鼠(whole-body homozygous miR-137 knockout mice)的觀察中，已知Knockout (miR-137-/-)小鼠大約在出生後21天死亡，且有嚴重的生長遲緩現象，在隨機血糖的測量上，Wild-type (miR-137+/+)、Heterozygous (miR-137+/-)、Knockout小鼠呈現高中低的趨勢，透過組織染色發現Knockout小鼠的脂肪細胞嚴重萎縮(Adipose atrophy)。根據上述的初步結果，我們認為miR-137可能與小鼠的生理代謝有關，而由於Knockout小鼠早夭的特性，本次研究是以Heterozygous老鼠為主，以一般飼料（Regular diet; RD）飼養到七周大成鼠時，依據實驗設計再給予特定組別高油脂飼料（High-fat diet; HFD），是一種常見在活體實驗中給予代謝壓力的實驗策略。首先在全程餵食一般飼料的組別中，Heterozygous小鼠體重上升的較多，但攝食量並沒有顯著差異，血清中的三酸甘油脂和肝臟中的脂肪酸氧化(β-oxidation)的決定速率酵素Cpt1a 表現量則比起控制組更高，表示miR-137的減少確實對於小鼠的代謝產生了影響。進一步我們給予小鼠HFD，一開始在不限制HFD飲食量的組別中，Heterozygous小鼠體重上升的較多，但攝食量也比起控制組更高，暗示了miR-137和小鼠食慾的關係。而進一步針對小鼠的代謝，給予相同量的HFD (Pair-feeding)，發現Heterozygous小鼠的增重幅度並沒有如預期地和控制組有所不同，表示這個根據這個實驗策略，不同基因型（Genotype）小鼠的代謝並沒有差異。。 結果顯示，在活體中miR-137的減少除了影響小鼠的代謝，也影響了小鼠對於高油脂飼料的食慾，我們也期待miR-137能成為具有潛力的調控代謝及食慾的指標，甚至有朝一日能應用於代謝症候群的治療上，進而改善族群的健康。

MicroRNA is a class of non-coding RNA which is about 22 nucleotides long and has been widely studied in the regulation of gene expression. Recent studies revealed that miR-137 plays a potential role in neural development and cancer biology. However, the direct investigation in vivo for miR-137 is still rare. In the first, we had validated the top three miR-137 highly-expressed organs: brain, heart and liver. According to the accessible material in our lab, we chose doxorubicin, acetaminophen, lipopolysaccharide and high-fat diet as a short-term stress to affect miR-137 expression. Only high-fat diet feeding for 1 month downregulated the expression level of miR-137 in mouse liver. Besides, we had generated whole-body homozygous miR-137 knockout mice (miR-137-/-), and observed that miR-137-/- mice were died around 21-days-old with dramatic growth retardation. The random blood glucose was measured in a lower trend between the wild-type (miR-137+/+), heterozygous knockout (miR-137+/-) and miR-137-/- mice. In addition, the adipose tissue was severely atrophy in the miR-137-/- mice. We hypothesized that miR-137 would affect the physiological metabolism, and focused on miR-137+/- adult mice for our research. First, during regular diet-feeding, miR-137+/- mice gained much weight with no significantly different amount of food intake. Serum triglyceride and Cpt1a mRNA expression level were elevated in miR-137+/- mice compared to the wild-type control. Thus, miR-137 deficiency has impact on mouse basal metabolism. Furtherly, we subjected mice to a high-fat diet (HFD), which was commonly used as metabolism stress. With the ad libitum HFD-feeding, we found that miR-137+/- ate more HFD with heavier body weight compared to the wild-type control, linking miR-137 and appetite for the first time. We then introduced pair HFD-feeding to mice targeting for metabolism, but results indicated that there was no difference in metabolism between genotypes based on this strategy. In conclusion, miR-137 may play a role in both metabolism and appetite in vivo. We expect that miR-137 may be a potential marker for regulation of metabolism and appetite and a useful therapeutic target applied in the metabolic disease.