BODIPY-Cholesterol is cholesterol tagged with a boron dipyrromethene difluoride (BODIPY) fluorophore used for monitoring sterol uptake and inter-organelle sterol flux in cells with excitation of 480 nm and emission of 508 nm.[1] The excretion of BODIPY cholesterol from late endoplasmic organelles depends on acidic lipase and Niemann pickc1 protein.[6]
In vitro, treatment with Bodipy-cholesterol in cells, prominent PM labeling is observed at 2–5 min; however, upon ≥30 min incubations, it is also observed the fluorescence labeling of intracellular structures.[2] In vitro efficacy test it suggested that accumulation of BODIPY-cholesterol in the media gave more reproducible values than assaying for the loss of the compound from the cells.[3] With 1 μg BODIPY-cholesterol analogs show a similar cellular localization in HeLa cells and exhibit similar cholesterol efflux properties from THP1 cells to HDL acceptors.[4] BODIPY-cholesterol efflux clearly increased when treatment of fibroblasts with the Hh pathway agonist SAG, which enhances Ptc protein expression, or over-expression of human Ptc in yeast.[5] Treatment with miR-758 inhibitor obviously increased ABCA1-dependent cholesterol efflux by BODIPY-Cholesterol efflux assay.[7] Micrographs of EPCs incubated with HDL labeld bodipy-cholesterol (50 μg/ml) 30 min, small cytoplasmic vesicles as well as large positive MVBs containing intraluminal microvesicles, tightly-packed with reaction products could be displayed. The internalized-HDL-derived bodipy-cholesterol was also spread within many of the stacked Golgi cisterns and the TGN.[8]
参考文献:
[1].Wüstner D, et al. Potential of BODIPY-cholesterol for analysis of cholesterol transport and diffusion in living cells. Chem Phys Lipids. 2016 Jan;194:12-28.
[2].H?ltt?-Vuori M, et al. BODIPY-cholesterol: a new tool to visualize sterol trafficking in living cells and organisms. Traffic. 2008 Nov;9(11):1839-49.
[3].Sankaranarayanan S, et al. A sensitive assay for ABCA1-mediated cholesterol efflux using BODIPY-cholesterol. J Lipid Res. 2011 Dec;52(12):2332-2340.
[4].Liu Z, et al. Synthesis of cholesterol analogues bearing BODIPY fluorophores by Suzuki or Liebeskind-Srogl cross-coupling and evaluation of their potential for visualization of cholesterol pools. Chembiochem. 2014 Sep 22;15(14):2087-96.
[5].Bidet M, et al. The hedgehog receptor patched is involved in cholesterol transport. PLoS One. 2011;6(9):e23834.
[6].Kanerva K, et al. LDL cholesterol recycles to the plasma membrane via a Rab8a-Myosin5b-actin-dependent membrane transport route. Dev Cell. 2013 Nov 11;27(3):249-62.
[7].Yao Y, et al. Glucagon-like peptide-1 contributes to increases ABCA1 expression by downregulating miR-758 to regulate cholesterol homeostasis. Biochem Biophys Res Commun. 2018 Mar 4;497(2):652-658.
[8].Srisen K, et al. Human endothelial progenitor cells internalize high-density lipoprotein. PLoS One. 2013 Dec 30;8(12):e83189.
BODIPY-Cholesterol 是用硼二氟甲烷 (BODIPY) 荧光团标记的胆固醇，用于监测细胞中甾醇摄取和细胞器间甾醇通量，激发波长为 480 nm，发射波长为 508 nm。[1] 晚期内质细胞器排泄 BODIPY 胆固醇依赖于酸性脂肪酶和 Niemann pickc1 蛋白。[6]
在体外，在细胞中用 Bodipy-cholesterol 处理，在 2-5 分钟时观察到显着的 PM 标记；然而，在≥30分钟的孵育后，也观察到细胞内结构的荧光标记。[2]体外功效测试表明，BODIPY-胆固醇在培养基中的积累比测定具有更高的可重复性值[3]1 μg BODIPY-胆固醇类似物在 HeLa 细胞中表现出相似的细胞定位，并表现出相似的胆固醇从 THP1 细胞流出到 HDL 受体的特性。[4]当用 Hh 通路激动剂 SAG 处理成纤维细胞时，BODIPY-胆固醇流出明显增加，这会增强 Ptc 蛋白表达，或人 Ptc 在酵母中的过度表达。[5] 通过 BODIPY-Cholesterol 流出测定，用 miR-758 抑制剂处理明显增加了 ABCA1 依赖性胆固醇流出。[7] EPCs 显微照片与 HDL 标记的 bodipy-cholesterol (50 μg/ml) 孵育 30 分钟，小细胞质囊泡以及大的阳性 MVBs 含有内可以显示与反应产物紧密堆积的管腔微泡。内化 HDL 衍生的身体胆固醇也分布在许多堆叠的高尔基池和 TGN 中。[8]
| Cell experiment [1]: | |
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Cell lines |
Human primary fibroblasts |
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Preparation Method |
NPC1‐deficient GM3123 fibroblasts were incubated with Rhodamine‐dextran overnight to label terminal endocytic compartments, followed by incubation with 5 μg/mL (8.7 μm) BODIPY-Cholesterol in normal growth medium containing 5% fetal calf serum for 4 h, or 0.5 μm BODIPY-Cholesterol in medium containing 5% lipoprotein‐starved serum for 23 h. |
|
Reaction Conditions |
5 μg/mL, 4h |
|
Applications |
Time‐dependent partitioning of BODIPY-Cholesterol in the lysosomes of human primary fibroblasts. |
| Cell experiment [2]: | |
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Cell lines |
NIH 3T3 |
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Preparation Method |
NIH 3T3 were cultured in 24-well plates, incubated for 2 h with 2.5 μM BODIPY-cholesterol, and rinsed. Physiological buffer with or without 30 nM ShhN was added to the wells, and the BODIPY fluorescence intensity of the supernatants was measured after 1 h. |
|
Reaction Conditions |
2.5 μM; 2 h |
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Applications |
The BODIPY fluorescence measured in the supernatants after 1 h was significantly lower in wells where ShhN protein was added compared to wells that did not contain ShhN. |
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参考文献: [1]. H?ltt?-Vuori M, et al. Use of BODIPY-Cholesterol (TF-Chol) for Visualizing Lysosomal Cholesterol Accumulation. Traffic. 2016 Sep;17(9):1054-7. [2]. Bidet M, et al. The hedgehog receptor patched is involved in cholesterol transport. PLoS One. 2011;6(9):e23834. |
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