一水草酸钙晶体刺激肾近端小管细胞后miRNA表达谱的变化实验方法详情页

实验方法> RNA实验技术> RNA原位杂交>一水草酸钙晶体刺激肾近端小管细胞后miRNA表达谱的变化

一水草酸钙晶体刺激肾近端小管细胞后miRNA表达谱的变化

关键词： miRNA 表达谱芯片来源：伯豪生物

华中科技大学同济医学院泌尿外科研究所的研究人员证实，在一水草酸钙刺激肾近端小管细胞后，miRNA的表达谱发生了变化，这将暗示着miRNA也许在结石形成中发挥作用。背景介绍结石形成的原因至今尚未探知，而一水草酸钙（COM）是结石中主要成分，本文通过COM刺激HK-2细胞后的miRNA表达谱的变化，初步探讨miRNA在刺激过程中的作用。文章亮点通过LDH和DAPI染色方法测定最适宜浓度，通过miRNA和mRNA芯片测定表达谱变化，通过real-time PCR鉴定表达谱芯片，通过GO和PATHWAY分析预测生物学行为(课题相关Agilent human miRNA microarrays技术服务由上海伯豪生物技术有限公司提供)。结果分析1mmol/L的COM晶体24h刺激完后有25条miRNAs表达超过了1.5倍，其中16条是上升的，9条下降。 Dysregulated miRNAs after COM crystals exposure

Probe_ID	foldchange	Probe_ID	Foldchange
Up-regulated		Down-regulated
hsa-miR-371-5p	3.069941	hsa-miR-425*	0.069018
hsa-miR-3125	2.71054	hsv1-miR-H6-3p	0.071335
hsa-miR-638	2.682127	hsv1-miR-H7*	0.078529
hsa-miR-3195	2.251545	hsa-miR-933	0.09623093
ebv-miR-BART13	2.155447	hsa-miR-191*	0.303036
hsa-miR-135a*	2.078597	hsa-let-7f-1*	0.366442
hsa-miR-1274b	2.004063	hsa-miR-4284	0.393521
hsa-miR-497	1.918096	hsa-miR-4313	0.485941
hsa-miR-1181	1.829789	hsa-let-7b*	0.48814
hsa-miR-195	1.706755
hsa-miR-1260b	1.693521
hsa-miR-21*	1.688719
hsa-miR-642b	1.644018
hsa-miR-1260	1.585072
hsv1-miR-H8	1.532887
hsa-miR-1274a	1.505559

通过GO和PATHWAY分析这些变化的miRNA与细胞凋亡、代谢反应、信号传递等生物学效应有关, 提示miRNA在结石形成过程中可能存在作用。 Analysis of biological processes of the predicted miRNAs targets by GOstat

miRNAs	GO process	Target genes	Count	%	P-value
hsa-miR-497	GO:0043549:regulation of kinase activity	PPP2R1A, MADD, PTPLAD1, DRD5, PDCD4, CDC25A, CDC25B, CBLC, TARBP2, PRKAR2A, APP, TSPYL2, DUSP9, HTR2A	14	0.3688	0.04444
	GO:0007249:I-kappaB kinase/NF-kappaB cascade	PTPLAD1, SNIP1, TLR1, IKBKB, TNIP2	5	0.1317	0.04600
	GO:0031980:mitochondrial lumen	GLS2, MRPL40, PDPR, FECH, HARS2, TMLHE, CARS2, YARS2, AK3L1, LRPPRC, KARS	11	0.2898	0.02395
	GO:0005739:mitochondrion	MRPL40, ACOX1, D2HGDH, CCDC19, NLRX1, AK3L1, KCNJ11, KARS, GLS2, GPX1, GSR, HTRA2, CISD3, SLC25A22, HARS2, SLC25A2, BDH2, AATK, PDPR, PPP2R1A, TRMU, FECH, NOL3, MRPS2, MMAB, RAB40AL, UCP2, TMLHE, SLC25A35, CARS2, YARS2, ALKBH3, LRPPRC	33	0.8693	0.03916
hsa-miR-638	GO:0009611:response to wounding	CCL11, MYF6, GPX1, TNFRSF1B, F10, MGLL, HOXB13, IL11	8	0.9335	0.00599
	GO:0010740:positive regulation of protein kinase cascade	GPX1, F10, BST2, IL11	4	0.4667	0.03180
	GO:0031099:regeneration	MYF6, GPX1, NR4A3	3	0.3501	0.03308
hsa-miR-1260	GO:0051252:regulation of RNA metabolic process	ZNF583, SNIP1, PDX1, ZNF253, SKAP1, IL11, APP, GFI1B, RAB26, SLA2, OTX1, ARID5A, SOX12, HDAC10, FOSB, NPAS4, SPEN, UBN1, ZNF192, ZNF786, MED8, SERBP1, TGIF2, CUX1, TFAP2E, MAPRE3, MED1, CARHSP1, FOXM1, HOXB13, FHL2, ZNF367, NEDD8, ZNF177, SUFU, ZNF615, ZNF169, TNFRSF1B, TSC22D2, TCEA3, HEXIM2, LHX1, OVOL1, GATAD2B, ZNF426, ZNF701, FOXD4, SIM2, CHD3, MYF6, ESRRA, ZNF565, CREBZF, TAF8, NR4A1, ZNF221, NR4A3, ZNF763, RFX3, MESP2, NCOR2	61	1.5204	0.00323
	GO:0045449:regulation of transcription	ZNF583, SNIP1, PDX1, ZNF253, PDCD4, CBX7, SKAP1, IL11, ARHGAP22, BZW1, APP, LBH, GFI1B, MED29, RAB26, SSBP2, POGZ, OTX1, SLA2, SOX12, ARID5A, HDAC10, FOSB, HDAC11, SPEN, NPAS4, UBN1, CCNL2, ZNF192, CCR6, PRDM4, ZNF786, MED8, PARP14, ZNF384, ERN2, TGIF2, CUX1, TFAP2E, MAPRE3, MED1, CARHSP1, FOXM1, TRIB3, HOXB13, FHL2, ZNF367, NEDD8, ZNF177, ZNF615, SUFU, ZNF169, TSC22D2, TCEA3, HEXIM2, LHX1, OVOL1, GATAD2B, ZNF426, RNF10, ZNF701, FOXD4, CHD3, SIM2, MYF6, SSRP1, ESRRA, ZBTB46, ZNF565, CREBZF, TAF8, NR4A1, ZNF221, NR4A3, ZNF629, PHF19, ZNF763, RFX3, IKBKB, MESP2, NCOR2	80	1.9940	0.00686
	GO:0043167:ion binding	SLC13A5, ZNF583, ZNRF1, ZNF253, TTL, APOBEC3C, APOBEC3D, MAP3K5, APP, CISD3, CDH23, RET, CAPNS2, FECH, PDXK, CRTAC1, BSN, RNASEH2A, WDR92, ZNF192, PRDM4, ZNF786, EIF2S2, ZNF384, UNC13A, NEK3, ZNF615, KCNJ1, RPS27, ZNF428, LHX1, OVOL1, RNF168, RNF10, ZNF426, STK19, GABRP, GIT1, NPLOC4, ESRRA, ZBTB46, B4GALT2, SETDB2, NR4A1, ZNF221, NR4A3, ACACB, ATP13A3, CYB561, PRPSAP2, ZNF629, TRIM56, KCNJ9, TMLHE, ZNF385D, SPTA1, TRAFD1, GLRA1, MLPH, GLRA3, CYP11B2, PDIA4, KCNJ11, KCNIP1, USP19, SMAP2, GFI1B, STAC2, TRIM9, DBR1, NSF, NT5C, POGZ, MICAL3, RPH3AL, CYB5B, ARL3, MYRIP, MAP3K15, ZNF691, TAF15, ERN2, CA4, CLCNKA, ZNF367, FHL2, EGLN2, ZNF177, BEST4, ZNF169, CYB561D1, TCEA3, CALML3, MMACHC, PRRG2, RASGRP2, DTNB, ENO2, MT1E, GATAD2B, GALNT11, ZNF701, CHD3, MT1M, NOS1, ZNF565, TRIM29, TRIM25, CYP4F8, RPS6KA3, FBLN1, RASSF5, PHF19, CDH15, ARSA, SCN4B, SUMF2, ZNF763, ALKBH3, RNF40	120	2.9910	0.03602
hsa-miR-1260b	GO:0051252:regulation of RNA metabolic process	ZNF583, SNIP1, PDX1, ZNF253, SKAP1, IL11, APP, GFI1B, RAB26, SLA2, OTX1, ARID5A, SOX12, HDAC10, FOSB, NPAS4, SPEN, UBN1, ZNF192, ZNF786, MED8, SERBP1, TGIF2, CUX1, TFAP2E, MAPRE3, MED1, CARHSP1, FOXM1, HOXB13, FHL2, ZNF367, NEDD8, ZNF177, SUFU, ZNF615, ZNF169, TNFRSF1B, TSC22D2, TCEA3, HEXIM2, LHX1, OVOL1, GATAD2B, ZNF426, ZNF701, FOXD4, SIM2, CHD3, MYF6, ESRRA, ZNF565, CREBZF, TAF8, NR4A1, ZNF221, NR4A3, ZNF763, RFX3, MESP2, NCOR2	61	1.5204	0.00323
	GO:0007242:intracellular signaling cascade	FGFR1, CARHSP1, RAB9A, RAB3D, PTPLAD1, DRD5, SNIP1, MAPKAPK3, FHL2, EGLN2, PDX1, RAB42, MAP3K5, DIRAS1, PRKAR2A, KISS1R, STAC2, RASGRP2, RAB26, TNIP2, LTB, GDI1, RET, C5AR1, PIK3C2B, SLA2, STMN4, NFKBIL1, DEPDC5, ARL3, RASL11B, RASSF5, RPS6KA3, RAB37, CNIH3, BBC3, SGSM3, RAB17, ASB1, IKBKB, UNC13A, MED1	42	1.0469	0.01837
hsa-miR-3125	GO:0006350:transcription	RNASEK, FOXM1, CBX4, TTLL5, ZNF367, ZNF177, CBX7, ZNF615, DPF1, ZNF169, GFI1B, OVOL1, GATAD2B, TWIST2, FOXD4, CHD3, ZNF548, ZBTB46, BRF2, TRIM29, SOX12, CDK9, DACH1, NR4A3, SPEN, HDAC11, HMGA1, NRIP2, CCR6, PRDM4, MED8, PARP14, ZNF384, RFX3, LRPPRC, PUF60, MED1	37	1.9004	0.01354
hsa-miR-3195	GO:0002064:epithelial cell development	AGPAT6, HOXB13	2	0.9009	0.02456
	GO:0004672:protein kinase activity	FGFR1, LMTK2, CDK9, IKBKB	4	1.8018	0.04223
hsa-miR-4284	GO:0008629:induction of apoptosis by intracellular signals	CUL3, CUL2, BBC3, CIDEA	4	0.1980	0.02512
hsa-miR-4313	GO:0045449:regulation of transcription	CARHSP1, ZNF583, GDF7, HOXB13, CBX7, PDCD4, IL11, DPF1, CCNE1, TSPYL2, MED29, OVOL1, TWIST2, ESRRA, ZBTB46, POGZ, BRF2, ARID5A, HDAC11, DACH1, ZNF497, HMGA1, CCNL2, ZNF629, TARBP2, PRDM4, MED8, PARP14, ZNF384, ZNF763, TGIF2, JAK3, CUX1, MAPRE3, MESP2, LRPPRC, NCOR2	37	2.2534	0.02350

原文出处Wang BH, Wu BL, Liu J, Yao WM, Xia D, Li L, Chen ZQ, Ye ZQ, Yu X. Analysis of Altered MicroRNA Expression Profiles in Proximal Renal Tubular Cells in Response to CalciumOxalate Monohydrate Crystal Adhesion: Implications for Kidney Stone Disease. Plos One.2014,9(7): e101306.

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