进行性半面萎缩症患者抽脂部位脂肪源性干细胞生物学特性的体外研究

In Vitro Study on the Biological Characteristics of Adipose-Derived Stem Cells from Liposuction Area in Patients with Progressive Hemifacial Atrophy.

作者信息

Gao Qiuni, Qi Zuoliang, Jin Xiaolei, Yang Zhenyu, Yuan Xihang, Hu Yuling, Yang Xiaonan

机构信息

Plastic Surgery Department, Plastic Surgery Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, No.33 Badachu Road, Shijingshan District, Beijing, 100144, China.

出版信息

Aesthetic Plast Surg. 2022 Oct;46(5):2526-2538. doi: 10.1007/s00266-022-02902-x. Epub 2022 May 12.

DOI:10.1007/s00266-022-02902-x

PMID:35552479

Abstract

OBJECTIVE

In this study, we investigate the biological characteristics of ADSCs from the liposuction area in patients with hemifacial atrophy in vitro.

METHODS

ADSCs were respectively extracted from the donor site of patients with hemifacial atrophy and healthy ones. ADSCs of two groups were respectively tested for proliferation ability, phenotype, multipotency, migration ability, self-repair ability, apoptosis, and autophagy. Exosomes extracted from the supernatant of two groups were detected by NTA particle size, electron microscopy (TEM), and WB for CD63 and TSG10, respectively.

RESULTS

CCK-8 showed a statistically less increase in cell proliferation in PHA-ADSCs after the sixth day. ADSCs in both groups had typical phenotypes and multidirectional abilities. PHA-ADSCs exhibited weaker droplet formation. The cell migration ability in PHA-ADSCs was weaker tested by Transwell assay. The live/dead proportion calculated by ImageJ following calcein-AM/PI double staining revealed live cells in PHA-ADSCs was 46.11% compared with 54.21% in NORM-ADSCs after OGD treatment. A significant down-regulation of ATG7 and ATG12 and a higher percentage of apoptosis were found in PHA-ADSCs. A significant up-regulation of BAX occurred in PHA-ADSCs.ARPC5 expression in the PHA group was extremely distinct down-regulated.CDKN1A and CDKN2A expression in the PHA group was significantly up-regulated.WB analyses confirmed that both groups' ADSCs-Exosomes surface markers CD63 and TSG101 were positively expressed but varied significantly.

CONCLUSIONS

PHA-ADSCs exhibited a poorer proliferation ability, higher apoptosis percentage, weaker lipid droplets formation, weaker cell migration, poorer intolerance to OGD, aging earlier, and weaker self-renewal and repairability.PHA-ADSCs-Exosomes showed low expressions of CD63 and TSG101.This study provides strong evidence that the addition of exosomes with specific cytokines can improve the fat survival rate after fat filling in patients with hemifacial atrophy.

NO LEVEL ASSIGNED

This journal requires that authors 42 assign a level of evidence to each submission to which 43 Evidence-Based Medicine rankings are applicable. This 44 excludes Review Articles, Book Reviews, and manuscripts 45 that concern Basic Science, Animal Studies, Cadaver 46 Studies, and Experimental Studies. For a full description of 47 these Evidence-Based Medicine ratings, please refer to the 48 Table of Contents or the online Instructions to Authors 49 https://www.springer.com/00266 .

摘要

目的

在本研究中，我们在体外研究半侧面部萎缩患者抽脂部位脂肪间充质干细胞（ADSCs）的生物学特性。

方法

分别从半侧面部萎缩患者及健康者的供区提取ADSCs。对两组ADSCs分别进行增殖能力、表型、多能性、迁移能力、自我修复能力、凋亡及自噬检测。分别采用NTA粒径分析、电子显微镜（TEM）及蛋白质免疫印迹法（WB）检测两组上清液中提取的外泌体的CD63和TSG10。

结果

CCK-8检测显示，第6天后PHA-ADSCs细胞增殖的增加在统计学上较少。两组ADSCs均具有典型表型和多向分化能力。PHA-ADSCs形成脂滴的能力较弱。通过Transwell实验检测，PHA-ADSCs的细胞迁移能力较弱。钙黄绿素-AM/PI双染后用ImageJ计算的活/死比例显示，OGD处理后PHA-ADSCs中的活细胞为46.11%，而NORM-ADSCs中的活细胞为54.21%。PHA-ADSCs中发现ATG7和ATG12显著下调，凋亡百分比更高。PHA-ADSCs中BAX显著上调。PHA组中ARPC5表达极度下调。PHA组中CDKN1A和CDKN2A表达显著上调。WB分析证实，两组ADSCs-Exosomes表面标志物CD63和TSG101均呈阳性表达，但差异显著。

结论

PHA-ADSCs表现出较差的增殖能力、较高的凋亡百分比、较弱的脂滴形成能力、较弱的细胞迁移能力、对OGD的耐受性较差、衰老较早以及较弱的自我更新和修复能力。PHA-ADSCs-Exosomes显示CD63和TSG101低表达。本研究提供了有力证据，即添加含有特定细胞因子的外泌体可提高半侧面部萎缩患者脂肪填充后的脂肪存活率。