A microRNA cluster controls fat cell differentiation and adipose tissue expansion by regulating SNCG

We present here lipidomic data from subcutaneous adipose tissue from 20 miR-KO mice and Wt controls under normal chow (1:1, male/females) (5 animals/experimental condition). Prior to lipid extraction, fat samples were spiked with internal standards and solvent of standards was removed by vacuum centrifugation. Then, samples were homogenized with a Precellys® 24 tissue homogenizer (Bertin Instruments). Homogenates containing a wet weight of 2 mg were extracted according to the protocol described by Bligh and Dyer. The analysis of lipids was performed by direct flow injection analysis (FIA), using either a triple quadrupole mass spectrometer (QQQ; FIA-MS/MS) or a hybrid quadrupole-Orbitrap high-resolution mass spectrometer (FIA-FTMS). Both instruments were equipped with a heated electrospray ionization source. FIA-MS/MS (QQQ) was performed in positive ion mode. The following neutral losses were applied: PE, 141 and PI, 277. PE-based plasmalogens (PE P) were analyzed according to the principles described by Berry and colleagues. Sphingosine-based Cer were analyzed using a fragment ion of <i>m/z</i> 264. Correction for isotopic overlap of lipid species was performed for all lipid classes. TG and DG were recorded in positive ion mode as [M+NH₄]⁺ in <i>m/z</i> range 500–1000 and a target resolution of 140,000 (at <i>m/z</i> 200). PC and SM were analyzed as [M+HCOO]^- in negative ion mode in <i>m/z</i> range 520–960 at the same resolution setting. Peak assignment was performed with the ALEX software and applied a mass accuracy of less than 3 ppm. Quantification was achieved by multiplication of the spiked-in IS amount with the analyte-to-IS intensity ratio. The following lipid species were applied as internal standards: Cer 18:1;O2/14:0, Cer 18:1;O2/17:0, DG 14:0/14:0/0:0, DG 20:0/20:0/0:0, HexCer 18:1;O2/12:0, HexCer 18:1;O2/17:0, LPC 13:0/0:0, LPC 19:0/0:0, PC 14:0/14:0, PC 22:0/22:0, PE 14:0/14:0, PE 20:0/20:0, PI 17:0/17:0, SM 18:1;O2/12:0, TG 17:0/17:0/17:0, and TG 19:0/19:0/19:0. Extracted data were processed by self-programmed Macros.

本研究公开了20只miRNA敲除（miR-KO）小鼠与野生型（wild type, Wt）对照小鼠的皮下脂肪组织脂质组学数据。所有受试动物均饲喂正常饲料，雌雄比例为1:1，每个实验条件下设5只动物。
脂质提取前，脂肪样本已加入内标，随后通过真空离心去除内标溶剂。
随后使用Precellys®24组织匀浆仪（伯特林仪器公司，Bertin Instruments）对样本进行匀浆处理。称取湿重2 mg的匀浆物，按照Bligh与Dyer建立的经典方法完成脂质提取。
脂质分析采用直接流动注射分析（direct flow injection analysis, FIA），分别使用三重四极杆质谱仪（triple quadrupole mass spectrometer, QQQ；FIA-MS/MS）与四极杆-轨道阱混合高分辨质谱仪（hybrid quadrupole-Orbitrap high-resolution mass spectrometer, FIA-FTMS）完成检测。两台仪器均配备加热电喷雾电离源。
FIA-MS/MS（QQQ）采用正离子模式检测，设置的中性丢失扫描参数为：磷脂酰乙醇胺（phosphatidylethanolamine, PE）对应141，磷脂酰肌醇（phosphatidylinositol, PI）对应277。磷脂酰乙醇胺缩醛磷脂（PE P）的分析参照Berry及其团队提出的方法原理进行。以神经鞘氨醇为骨架的神经酰胺（ceramide, Cer）通过检测质荷比（m/z）264的特征碎片离子完成定性分析。
针对各脂质类别，均完成了同位素峰重叠校正。三酰甘油（triacylglycerol, TG）与二酰甘油（diacylglycerol, DG）采用正离子模式，以[M+NH₄]⁺为检测离子，扫描质荷比范围为500–1000，目标分辨率设为140000（在m/z 200处）。磷脂酰胆碱（phosphatidylcholine, PC）与鞘磷脂（sphingomyelin, SM）采用负离子模式，以[M+HCOO]⁻为检测离子，扫描质荷比范围为520–960，分辨率设置与前述一致。
峰指认工作通过ALEX软件完成，设置的质量准确度阈值低于3 ppm。定量分析通过将加入的内标量与分析物/内标信号强度比值相乘得到最终结果。
本次实验使用的内标脂质包括：Cer 18:1;O2/14:0、Cer 18:1;O2/17:0、DG 14:0/14:0/0:0、DG 20:0/20:0/0:0、己糖基神经酰胺（hexosylceramide, HexCer）18:1;O2/12:0、HexCer 18:1;O2/17:0、溶血磷脂酰胆碱（lysophosphatidylcholine, LPC）13:0/0:0、LPC 19:0/0:0、PC 14:0/14:0、PC 22:0/22:0、PE 14:0/14:0、PE 20:0/20:0、PI 17:0/17:0、SM 18:1;O2/12:0、TG 17:0/17:0/17:0以及TG 19:0/19:0/19:0。
提取得到的原始数据通过自行编写的宏程序进行处理。