Gao Shuping, Calcagni Maurizio, Welti Manfred, Hemmi Sonja, Hild Nora, Stark Wendelin J, Bürgisser Gabriella Meier, Wanner Guido A, Cinelli Paolo, Buschmann Johanna
Division of Trauma Surgery, University Hospital Zurich, ZKF, Sternwartstrasse 14, CH-8091 Zurich, Switzerland.
Division of Plastic and Hand Surgery, University Hospital Zurich, ZKF, Sternwartstrasse 14, CH-8091 Zurich, Switzerland.
Injury. 2014 Jun;45(6):974-80. doi: 10.1016/j.injury.2014.02.035. Epub 2014 Mar 12.
Fractures with a critical size bone defect are associated with high rates of delayed- and non-union. The treatment of such complications remains a serious issue in orthopaedic surgery. Adipose derived stem cells (ASCs) combined with biomimetic materials can potentially be used to increase fracture healing. Nevertheless, a number of requirements have to be fulfilled; in particular, the insufficient vascularisation of the bone constructs. Here, the objectives were to study the impact of ASC-derived osteoblasts on ASC-derived endothelial cells in a 3D co-culture and the effect of 40wt% of amorphous calcium phosphate nanoparticles on the proliferation and differentiation of ASC-derived endothelial cells when present in PLGA.
Five primary ASC lines were differentiated towards osteoblasts (OBs) and endothelial cells (ECs) and two of them were chosen based on quantitative PCR results. Either a mono-culture of ASC-derived EC or a co-culture of ASC-derived EC with ASC-derived OB (1:1) was seeded on an electrospun nanocomposite of poly-(lactic-co-glycolic acid) and amorphous calcium phosphate nanoparticles (PLGA/a-CaP; reference: PLGA). The proliferation behaviour was determined histomorphometrically in different zones and the expression of von Willebrand Factor (vWF) was quantified.
Independently of the fat source (biologic variability), ASC-derived osteoblasts decelerated the proliferation behaviour of ASC-derived endothelial cells in the co-culture compared to the mono-culture. However, expression of vWF was clearly stronger in the co-culture, indicating further differentiation of the ASC-derived EC into the EC lineage. Moreover, the presence of a-CaP nanoparticles in the scaffold slowed the proliferation behaviour of the co-culture cells, too, going along with a further differentiation of the ASC-derived OB, when compared to pure PLGA scaffolds.
This study revealed significant findings for bone tissue-engineering. Co-cultures of ASC-derived EC and ASC-derived OB stimulate each other's further differentiation. A nanocomposite with a-CaP nanoparticles offers higher mechanical stability, bioactivity and osteoconductivity compared to mere PLGA and can easily be seeded with pre-differentiated EC and OB.
具有临界尺寸骨缺损的骨折与骨不连和延迟愈合的高发生率相关。此类并发症的治疗仍是骨科手术中的一个严重问题。脂肪来源干细胞（ASC）与仿生材料联合使用可能有助于促进骨折愈合。然而，必须满足许多要求；特别是骨构建物血管化不足的问题。在此，本研究的目的是研究三维共培养中ASC来源的成骨细胞对ASC来源的内皮细胞的影响，以及当聚乳酸-羟基乙酸共聚物（PLGA）中存在40wt%的无定形磷酸钙纳米颗粒时，其对ASC来源的内皮细胞增殖和分化的影响。
将5个原代ASC系分别诱导分化为成骨细胞（OB）和内皮细胞（EC），并根据定量PCR结果选择其中2个细胞系。将ASC来源的EC进行单培养或将ASC来源的EC与ASC来源的OB按1:1比例进行共培养，接种于聚乳酸-羟基乙酸共聚物与无定形磷酸钙纳米颗粒的电纺纳米复合材料（PLGA/a-CaP；对照：PLGA）上。通过组织形态计量学方法测定不同区域的增殖行为，并对血管性血友病因子（vWF）的表达进行定量分析。
与单培养相比，无论脂肪来源如何（生物变异性），在共培养中ASC来源的成骨细胞均会减缓ASC来源的内皮细胞的增殖行为。然而，共培养中vWF的表达明显更强，表明ASC来源的EC进一步分化为内皮细胞谱系。此外，与纯PLGA支架相比，支架中a-CaP纳米颗粒的存在也减缓了共培养细胞的增殖行为，同时伴随着ASC来源的OB的进一步分化。
本研究为骨组织工程提供了重要发现。ASC来源的EC与ASC来源的OB共培养可相互促进进一步分化。与单纯的PLGA相比，含有a-CaP纳米颗粒的纳米复合材料具有更高的机械稳定性、生物活性和骨传导性，并且可以很容易地接种预分化的EC和OB。
