Analysis of circRNA and mRNA expression profiles in Alzheimer's disease mice treated with islet amyloid polypeptide
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摘要:
目的 探究胰岛淀粉样多肽(IAPP)治疗阿尔茨海默病(AD)小鼠的脑组织环状RNA(circRNA)和信使RNA(mRNA)表达谱变化及分子机制。 方法 将6只APP/PS1小鼠按随机数字表法随机分为IAPP干预组和参照组，每组3只。干预组小鼠腹腔注射0.5 μmol/L的IAPP(200 μg/kg), 参照组小鼠注射相同剂量的磷酸缓冲盐溶液。使用基因芯片技术检测2组小鼠之间差异表达的circRNAs和mRNAs，通过qRT-PCR对芯片结果进行验证，并对差异表达的mRNAs进行GO和KEGG通路分析。 结果 按照差异倍数的绝对值(|FC|)≥1.5且P < 0.05的筛选标准, IAPP干预组与参照组之间有899个差异表达circRNAs，343个上调和556个下调，差异表达mRNAs有2 257个，1 310个上调和947个下调。qRT-PCR验证结果均与芯片结果一致。GO分析显示，差异表达mRNAs主要富集于G蛋白偶联受体、细胞代谢等生物功能；KEGG通路分析显示，差异表达mRNAs主要与柠檬酸循环、蛋白酶体、IL-17和趋化因子信号通路等相关。 结论 腹腔注射IAPP后AD小鼠脑组织中circRNAs和mRNAs表达谱发生显著变化，提示这些差异表达的circRNAs和mRNAs可能是IAPP治疗AD小鼠的关键机制。 Abstract:Objective To investigate the changes of circulating RNA (circRNA) and messenger RNA (mRNA) expression profiles in brain tissue of mice with Alzheimer's disease (AD) treated with islet amyloid polypeptide (IAPP) and the molecular mechanisms. Methods According to the number table method, 6 APP/PS1 mice were randomly divided into IAPP intervention group and reference group, 3 mice in each group. The intervention group mice were injected intraperitoneally with 0.5 μmol/L of IAPP (200 μg/kg), the reference group mice were injected with the same dose of phosphate buffer saline. Differentially expressed circRNAs and mRNAs between the two groups of mice were detected using gene chip technology. Chip results were validated by qRT-PCR. Differentially expressed mRNAs were analyzed by GO and KEGG pathways. Results According to the screening criteria of difference fold change (|FC|) ≥1.5 and P < 0.05, there were 899 differentially expressed circRNAs, 343 up-regulated and 556 down-regulated, and 2 257 differentially expressed mRNAs, 1 310 up-regulated and 947 down-regulated, between the IAPP intervention group and the reference group. qRT-PCR validation results were consistent with the gene chip results. GO analysis showed that the differentially expressed mRNAs were mainly enriched in G protein-coupled receptor, cell metabolism and other biological functions. KEGG pathway analysis showed that they were mainly involved in citrate cycle, proteasome, IL-17 and chemokine signaling pathway, etc. Conclusion The expression profiles of circRNAs and mRNAs in the brain tissue of AD mice were significantly changed after intraperitoneal injection of IAPP, suggesting that these differentially expressed circRNAs and mRNAs may be the key mechanisms of IAPP treatment in AD mice. -
Key words:
- Islet amyloid polypeptide /
- Alzheimer's disease /
- Circular RNA /
- Messenger RNA
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图 1 显著差异表达的mRNAs的KEGG通路富集分析
注：A为显著表达上调的mRNAs富集通路；B为显著表达下调的mRNAs富集通路。
Figure 1. KEGG pathway enrichment analysis of significantly differentially expressed mRNAs
表 1 目的基因引物名称、序列和长度
Table 1. Primer names, sequences and lengths of target genes
基因名称 双向引物序列 产物长度
(bp)mmu_circRNA_35991 正向: 5’-GGGGAACATTTTATCAAGGCT-3’ 119 反向: 5’-TGAACTTTTACAAGGTCCAACG-3’ mmu_circRNA_39224 正向: 5’-ACCAAACTCATTGATTTCCTGC-3’ 83 反向: 5’-GCTTGCTCCAAACTTCGCTT-3’ mmu_circRNA_43643 正向: 5’-CGAAGGCATGGAGACCTGTAA-3’ 89 反向: 5’-GCAGCCTGTCATCTTCAATCA-3’ mmu_circRNA_25778 正向: 5’-TGTTTGACCATCCAGACGCA-3’ 168 反向: 5’-CAGGTTACAGTTGCTGACCAGTG-3’ GAPDH 正向: 5’-CACTGAGCAAGAGAGGCCCTAT-3’ 144 反向: 5’-GCAGCGAACTTTATTGATGGTATT-3’ 
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表 2 IAPP干预组与参照组之间差异表达变化最显著的前10个circRNAs
Table 2. The top 10 circRNAs with the most significant differential expression changes between IAPP intervention group and control group
circRNA 表达 差异倍数 P值 基因名称 mmu_circRNA_35991 上调 3.94 0.049 Vav3 mmu_circRNA_21737 上调 3.89 0.002 Pde7b mmu_circRNA_39224 上调 3.84 0.021 Cit mmu_circRNA_44629 上调 3.52 0.032 Unc13c mmu_circRNA_014582 上调 2.99 0.035 Pard3 mmu_circRNA_007951 下调 3.92 0.041 Samd1 mmu_circRNA_38835 下调 2.93 0.001 Aasdh mmu_circRNA_29357 下调 2.75 0.002 Hic2 mmu_circRNA_33015 下调 2.70 0.001 4930515L03Rik mmu_circRNA_36800 下调 2.69 0.043 Fktn
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表 3 IAPP干预组与参照组之间差异表达变化最显著的前10个mRNAs
Table 3. The top 10 mRNAs with the most significant differential expression changes between IAPP intervention group and control group
mRNA 表达 差异倍数 P值 基因名称 ENSMUST00000060125 上调 14.39 0.022 Scn4b ENSMUST00000036172 上调 8.54 0.015 Sgpp2 ENSMUST00000111332 上调 6.94 0.007 Pcp4l1 ENSMUST00000071093 上调 5.71 0.017 Rims3 ENSMUST00000084650 上调 5.67 0.022 Gpr139 ENSMUST00000089200 下调 77.76 0.002 Cst7 ENSMUST00000040002 下调 14.17 0.049 Prr32 ENSMUST00000173014 下调 11.79 0.006 Dio3 ENSMUST00000001008 下调 9.29 0.001 Ccl3 ENSMUST00000040655 下调 7.36 0.012 H2-Aa
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表 4 IAPP干预组与参照组之间目的基因相对表达量水平(x±s)
Table 4. Relative expression levels of target genes between IAPP intervention group and control group (x±s)
组别 n mmu_circRNA_35991 mmu_circRNA_39224 mmu_circRNA_43643 mmu_circRNA_25778 IAPP干预组 3 25.23±8.33 12.63±0.50 4.67±1.18 0.30±0.10 参照组 3 2.88±0.92 4.54±0.59 1.79±0.28 0.93±0.17 t值 -4.626 -18.151 -4.108 5.686 P值 0.010 < 0.001 0.015 0.005
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