巨乌贼皮来源的凋亡模拟物对核因子 κB 受体激活剂配体/巨噬细胞集落刺激因子诱导的 RAW264.7 细胞破骨细胞生成的抑制作用。

Suppressive effects of an apoptotic mimicry prepared from jumbo-flying squid-skin phospholipids on the osteoclastogenesis in receptor activator of nuclear factor kappa B ligand/macrophage colony-stimulating factor-induced RAW 264.7 cells.

机构信息

Department of Animal Science and Technology, National Taiwan University, Taipei, Taiwan, ROC.

Seafood Technology Division, Fisheries Research Institute, Council of Agriculture, Executive Yuan, Keelung, Taiwan, ROC.

出版信息

J Chin Med Assoc. 2021 Jan 1;84(1):51-60. doi: 10.1097/JCMA.0000000000000447.

DOI:10.1097/JCMA.0000000000000447

PMID:33177403

Abstract

BACKGROUND

Liposomes containing docosahexaenoic acid (DHA) and phosphatidylserine were claimed to inhibit osteoclast formation and bone resorption in the inflammatory status. Herein, we proposed that an apoptotic mimicry (SQ liposome) prepared from squid-skin phospholipids can explore the suppressive osteoclastogenesis.

METHODS

The intermolecular fatty-acid composition in the phospholipid of squid-skin extract was analyzed by GC-FID. The SQ liposome structure was characterized by size distribution and zeta potential (ζ). RAW 264.7 cell is used to study the effect of SQ liposomes on osteoclast differentiation. Secretion of prostaglandin E2 (PGE2) and transforming growth factor-β (TGF-β) from RAW 264.7 cells were assayed. Antiosteoclastogenesis effects were performed via the tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cell (MNC) counting, bone resorption pit assay, and TRAP activity analysis. The specific gene expressions related to antiosteoclastogenesis were also detected.

RESULTS

An apoptotic mimicry through the use of a single-layer liposome (SQ liposome) with phosphatidylserine exposure contains DHA (28.7%) and eicosapentaenoic acid (EPA, 11.8%). Co-treatment with receptor activator of nuclear factor kappa B ligand (RANKL)/macrophage colony-stimulating factor induced RAW 264.7-cell differentiation into mature osteoclasts, thus enhancing PGE2 and TGF-β secretion. However, cotreatment with 1 mg/mL of SQ liposome restored (p < 0.05) the cell viabilities under the RANKL stress. Increased PGE2 levels was downregulated (p < 0.05) in cotreatments with 0.11 and 0.33 mg/mL of SQ liposome, but on the TGF-β levels were not (p > 0.05) influenced in SQ liposome cotreatments. Cotreatments with 0.33-1 mg/mL of SQ liposome suppressed (p < 0.05) the osteoclast maturation (such as decreased MNCs and bone pit formation), inhibited TRAP activities, and downregulated the osteoclastogenesis-related gene expressions.

CONCLUSION

In summary, current data support that a possible prevention of our prepared SQ liposomes which are rich in DHA and EPA on bone loss is through the suppression of osteoclastogenesis. Moreover, based on the results from this study an in vivo study warrants a further investigation.

摘要

背景

含有二十二碳六烯酸（DHA）和磷脂酰丝氨酸的脂质体被认为可以抑制炎症状态下破骨细胞的形成和骨吸收。在此，我们提出，从鱿鱼皮磷脂中制备的凋亡模拟物（SQ 脂质体）可以探索抑制破骨细胞生成的方法。

方法

通过 GC-FID 分析鱿鱼皮提取物中磷脂的分子间脂肪酸组成。通过粒径分布和 zeta 电位（ζ）对 SQ 脂质体结构进行表征。使用 RAW 264.7 细胞研究 SQ 脂质体对破骨细胞分化的影响。测定 RAW 264.7 细胞中前列腺素 E2（PGE2）和转化生长因子-β（TGF-β）的分泌。通过抗酒石酸酸性磷酸酶（TRAP）阳性多核细胞（MNC）计数、骨吸收陷窝试验和 TRAP 活性分析来评估抗破骨细胞生成作用。还检测了与抗破骨细胞生成相关的特定基因表达。

结果

通过使用含有 DHA（28.7%）和二十碳五烯酸（EPA，11.8%）的单层脂质体（SQ 脂质体）进行凋亡模拟，可产生类似凋亡的效果。核因子 kappa B 配体（RANKL）/巨噬细胞集落刺激因子共处理诱导 RAW 264.7 细胞分化为成熟破骨细胞，从而增强 PGE2 和 TGF-β 的分泌。然而，1mg/mL 的 SQ 脂质体共处理可恢复 RANKL 应激下的细胞活力（p<0.05）。在 0.11 和 0.33mg/mL 的 SQ 脂质体共处理中，升高的 PGE2 水平被下调（p<0.05），但 SQ 脂质体共处理对 TGF-β 水平没有影响（p>0.05）。0.33-1mg/mL 的 SQ 脂质体共处理可抑制（p<0.05）破骨细胞成熟（如 MNC 减少和骨陷窝形成）、抑制 TRAP 活性，并下调破骨细胞生成相关基因表达。