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Promoted Chondrogenesis of Cocultured Chondrocytes and Mesenchymal Stem Cells under Hypoxia Using In-situ Forming Degradable Hydrogel Scaffolds
Biomacromolecules ( IF 6.2 ) Pub Date : 2017-12-18 00:00:00 , DOI: 10.1021/acs.biomac.7b01271
Xiaobin Huang 1 , Yong Hou 2 , LeiLei Zhong 1 , Dechun Huang 3 , Hongliang Qian 3 , Marcel Karperien 1 , Wei Chen 2, 3
Affiliation  

We investigated the effects of different oxygen tension (21% and 2.5% O2) on the chondrogenesis of different cell systems cultured in pH-degradable PVA hydrogels, including human articular chondrocytes (hACs), human mesenchymal stem cells (hMSCs), and their cocultures with a hAC/hMSC ratio of 20/80. These hydrogels were prepared with vinyl ether acrylate-functionalized PVA (PVA-VEA) and thiolated PVA-VEA (PVA-VEA-SH) via Michael-type addition reaction. The rheology tests determined the gelation of the hydrogels was controlled within 2–7 min, dependent on the polymer concentrations. The different cell systems were cultured in the hydrogel scaffolds for 5 weeks, and the safranin O and GAG assay showed that hypoxia (2.5% O2) greatly promoted the cartilage matrix production with an order of hAC > hAC/hMSC > hMSC. The real time quantitative PCR (RT-PCR) revealed that the hMSC group exhibited the highest hypertrophic marker gene expression (COL10A1, ALPL, MMP13) as well as the dedifferentiated marker gene expression (COL1A1) under normoxia conditions (21% O2), while these expressions were greatly inhibited by coculturing with a 20% amount of hACs and significantly further repressed under hypoxia conditions, which was comparative to the sole hAC group. The enzyme-linked immunosorbent assay (ELISA) also showed that coculture of hMSC/hAC greatly reduced the catabolic gene expression of MMP1 and MMP3 compared with the hMSC group. It is obvious that the hypoxia conditions promoted the chondrogenesis of hMSC by adding a small amount of hACs, and also effectively inhibited their hypotrophy. We are convinced that coculture of hAC/hMSC using in situ forming hydrogel scaffolds is a promising approach to producing cell source for cartilage engineering without the huge needs of primary chondrocyte harvest and expansion.

中文翻译:

低氧条件下原位形成可降解水凝胶支架促进共培养的软骨细胞和间充质干细胞的软骨形成

我们研究了不同的氧气张力(21%和2.5%O 2)对在可pH降解的PVA水凝胶中培养的不同细胞系统的软骨形成的影响,这些凝胶包括人关节软骨细胞(hACs),人间充质干细胞(hMSCs)及其hAC / hMSC比为20/80的共培养物。这些水凝胶是通过迈克尔型加成反应用乙烯基醚丙烯酸酯官能化的PVA(PVA-VEA)和硫醇化的PVA-VEA(PVA-VEA-SH)制备的。流变学测试确定了水凝胶的凝胶化控制在2-7分钟内,具体取决于聚合物的浓度。在水凝胶支架中将不同的细胞系统培养5周,而番红素O和GAG分析表明缺氧(2.5%O 2)以hAC> hAC / hMSC> hMSC的顺序极大地促进了软骨基质的产生。实时定量PCR(RT-PCR)显示,在常氧条件下,hMSC组表现出最高的肥大标记基因表达(COL10A1,ALPL,MMP13)和去分化标记基因表达(COL1A1)(O 2为21%)),尽管与20%的hAC共同培养可大大抑制这些表达,并在缺氧条件下显着进一步抑制,这与唯一的hAC组相比。酶联免疫吸附试验(ELISA)还显示,与hMSC组相比,hMSC / hAC的共培养大大降低了MMP1和MMP3的分解代谢基因表达。显然,低氧条件通过添加少量的hAC促进了hMSC的软骨形成,并且还有效地抑制了它们的肥大。我们相信,使用原位形成水凝胶支架的hAC / hMSC共培养是生产软骨工程细胞源的有前途的方法,而无需原代软骨细胞的大量采集和扩增。
更新日期:2017-12-18
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