The effect of bioactive glass on adipose-derived msc osteogenesis in 3d cultures

Mohammad hossein karami Karami,1,* Mahboubeh kabiri,2 Mahshid heidari,3 Ali farazmand,4 Hana hanayi-ahvaz,5

1. Department of Biology, Cellular and molecular Biology, College of Science, University of Tehran,
2. Department of Biotechnology, College of Science, University of Tehran, Tehran
3. Department of Biotechnology, College of Science, University of Tehran
4. Department of Biology, Cellular and molecular Biology, College of Science, University of Tehran
5. Department of Stem Cell Biology, Stem Cell Technology Research center,

Abstract

Introduction

Regenerative medicine is an emerging field of biotechnology that combines various aspects of medicine, cell and molecular biology, materials science and bioengineering in order to regenerate, repair or replace tissues. bone regeneration is one of the focus points in the field of regenerative medicine because of huge number of patients needing bone surgery every year. adipose tissue (ad) represents a hot topic in regenerative medicine because of the tissue source abundance, the relatively easy retrieval, and the inherent biological properties of mesenchymal stem cells residing in its stroma .capturing the full potential of msc will likely require the development of novel in vitro culture techniques. here, we describe a novel and efficient system to culture mscs in 3d microaggregates composed of cells and bone mimicking inorganic nano-sized compounds and investigate their effects on msc osteogenesis.

Methods

 mscs were cultured in microwells (custom made from poly dimethylsiloxane (pdms)) to form 3d microaggregates in osteogenic induction medium. spheroids were generated using pdms-based concave micromolds developed using thin pdms membranes (figure1). in addition we addded ~0.42 μg bioactive glass into single multicellular aggregates formed from 80 mesenchymal stem. the concave microwells were coated with 5% pluronic acid to prevent cell attachment. cell aggregation and spheroid formation were observed daily under a microscope. calcium content assay was performed to evaluate osteogenesis. classic 2d cultures served as control group

Results

We successfully used concave microwell arrays to form uniform-sized ad derived mesenchymal stem cell spheroids. msc on cylindrical pdms microwells modified with pluronic acid formed homogenous 3d microaggregates. however some spheroids formed were joining together making larger spheroids also not stable in cylindrical microwells (figure2). our results showed that at day 7 and 14 post induction there was significantly greater calcium deposition in the microaggregates, relative to the 2d cell monolayer culture. in addition the resultant bio-glass endowed microaggregates cultured for two weeks under osteogenic induction medium showed prolonged formidability and rigidity. we would expect higher mechanical strength in composite endowed micro aggregates in comparison without bio glass or 2d controls.

Conclusion

 we developed concave microwells as a substrate for the formation of uniform-sized. in addition, we demonstrated that the concave microstructure offers a substantial advantage for the formation and harvesting of spheroids. bone mimicry micro tissues composed of both cells and bioactive glass can pave the way for macro tissue formation. we expect that our micro bone aggregates can be of so much use in the field of bone tissue engineering.

Keywords

Bioactive glass, adipose-derived msc, osteogenesis ,3d cultures