Lipidome modulation by dietary omega-3 polyunsaturated fatty acid supplementation or selective soluble epoxide hydrolase inhibition suppresses rough LPS-accelerated glomerulonephritis in lupus-prone mice

Lipopolysaccharide (LPS)-accelerated autoimmune glomerulonephritis (GN) in lupus-prone NZBWF1 mice is a preclinical model that is potentially applicable for investigating lipidome-modulating interventions. LPS can be expressed as one of two chemotypes: smooth LPS (S-LPS) and rough LPS (R-LPS) which is devoid of O-antigen polysaccharide sidechain. Since these chemotypes differentially affect TLR4-mediated immune cell responses, these differences may influence GN induction. Therefore, we initially compared the effects of subchronic i.p. injection for 5 wk with 1) Salmonella S-LPS, 2) Salmonella R-LPS, or 3) saline vehicle (VEH) (Study 1) in female NZBWF1 mice. R-LPS induced robust elevations in blood urea nitrogen, proteinuria, and hematuria that were not evident in VEH- or S-LPS-treated mice. Histopathologic examination of R-LPS-treated mice one week after final injection further revealed more robust hypertrophy, hyperplasia, thickened membranes, lymphocytic accumulation containing B and..., LC-MS/MS Quantitation of Plasma Oxylipins
Waters Oasis-HLB cartridges (part no. #WAT094226, lot no. #176A30323A) were used for sample preparation and clean-up purposes. Solid-phase extraction (SPE) cartridges were prepared for solid phase extraction by washing once with 2 ml of ethyl acetate, twice with 2 ml of methanol, and twice with 2 ml of 95:5 (v/v) water/methanol + 0.1% (v/v) acetic acid. Plasma was then loaded onto the cartridges, and samples were spiked with 10 μl of deuterated internal standard solution (16 nM BGB2-d4, 10 nM LTB4-d4, 16 nM 8,9-DiHETrE-d11, 16 nM 9-HODE-d4, 20 nM 15(S)-HETE-d8, 40 nM 5(S)-HETE-d8, 40 nM 8,9-EpETrE-d11) and 10 μl of antioxidant cocktail (0.2 mg/ml butylated hydroxytoluene, 0.2 mg/ml triphenylphosphine, 0.6 mg/ml EDTA). After loading samples, cartridges were washed with 1.5 ml of 95:5 (v/v) water/methanol + 0.1% (v/v) acetic acid. The SPE cartridges were dried with a low vacuum for about 20 minutes to pull out the water and other unwanted residues..., Microsoft Excel

脂多糖（Lipopolysaccharide, LPS）加速的狼疮易感NZBWF1小鼠自身免疫性肾小球肾炎（glomerulonephritis, GN）模型，是一种可用于探究脂质组调控干预手段的临床前模型。LPS可分为两种化学型：光滑型LPS（Smooth LPS, S-LPS）与缺乏O-抗原多糖侧链的粗糙型LPS（Rough LPS, R-LPS）。由于这两种化学型对Toll样受体4（TLR4）介导的免疫细胞应答存在差异化影响，其差异可能会影响肾小球肾炎的诱导过程。因此，本研究首先在雌性NZBWF1小鼠中对比了三种亚慢性腹腔注射方案持续5周的干预效果：1）鼠伤寒沙门氏菌光滑型LPS（Salmonella S-LPS）、2）鼠伤寒沙门氏菌粗糙型LPS（Salmonella R-LPS），以及3）生理盐水赋形剂（vehicle, VEH）（研究1）。

粗糙型LPS处理可显著升高小鼠的血尿素氮、蛋白尿与血尿水平，该变化未在赋形剂组或光滑型LPS处理组小鼠中观察到。末次注射后一周对粗糙型LPS处理组小鼠开展组织病理学检查，进一步发现其肾脏呈现更为显著的肥大、增生、膜增厚，以及包含B细胞与……的淋巴细胞聚集。血浆氧脂素（Plasma Oxylipins）的液相色谱-串联质谱（Liquid Chromatography-Tandem Mass Spectrometry, LC-MS/MS）定量分析：

采用沃特世（Waters）Oasis-HLB固相萃取小柱（货号：#WAT094226，批号：#176A30323A）完成样品制备与净化。固相萃取（Solid Phase Extraction, SPE）小柱需经如下活化步骤：以2 mL乙酸乙酯洗涤1次、2 mL甲醇洗涤2次，再以2 mL体积比为95:5的水/甲醇混合液（含0.1%体积比乙酸）洗涤2次。随后将血浆上样至小柱，并向样品中加入10 μL氘代内标溶液（含16 nM BGB2-d4、10 nM LTB4-d4、16 nM 8,9-二羟基二十碳三烯酸-d11（8,9-DiHETrE-d11）、16 nM 9-羟基十八碳二烯酸-d4（9-HODE-d4）、20 nM 15(S)-羟基二十碳四烯酸-d8（15(S)-HETE-d8）、40 nM 5(S)-羟基二十碳四烯酸-d8（5(S)-HETE-d8）与40 nM 8,9-环氧二十碳三烯酸-d11（8,9-EpETrE-d11））及10 μL抗氧化剂混合液（含0.2 mg/mL丁基羟基甲苯、0.2 mg/mL三苯基膦与0.6 mg/mL乙二胺四乙酸）。上样完成后，以1.5 mL体积比为95:5的水/甲醇混合液（含0.1%体积比乙酸）洗涤固相萃取小柱。随后将小柱置于低真空环境下干燥约20分钟，以去除水分及其他无关残留……微软Excel（Microsoft Excel）