Stem Cells Int. 2014;2014:576358.

Osteogenic Potential of Mouse Adipose-Derived Stem Cells Sorted for CD90 and CD105 in Vitro

Maiko Yamamoto1, Hidemi Nakata1, Jia Hao1, Joshua Chou2, Shohei Kasugai1, Shinji Kuroda1

1Oral Implantology and Regenerative Dental Medicine, Department of Oral Health Sciences, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan. fax/phone +81 3 5803 4656; 2Faculty of Science, University of Technology Sydney, P.O. Box 123, Broadway, Ultimo, Sydney, NSW 2007, Australia. fax/phone +61-2-9514-1709.

Contact:

Shinji Kuroda, D.D.S., Ph.D.

Oral Implantology and Regenerative Dental Medicine, Department of Oral Health Sciences, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan. Phone: +81-3-5803-4656, Fax: +81-3-5803-4656, E-mail: skuroda.mfc@tmd.ac.jp

 

Abstract

Adipose tissue-derived stromal cells, termed ASCs, play an important role in regenerative applications. They resemble mesenchymal stem cells owing to their inexhaustibility, general differentiation potential and plasticity and display a series of cell-specific and cluster-of-differentiation (CD) marker profiles similar to those of other somatic stem cells. Variations in phenotypes or differentiation are intimately associated with CD markers. The purpose of our study was to exhibit distinct populations of ASCs with differing characteristics for osteogenic differentiation. The primary cell batch of murine-derived ASCs was extracted from subcutaneous adipose tissue and the cells were sorted for the expression of the surface protein molecules CD90 and CD105 using flow cytometry. Each cell population sorted for CD90 and CD105 was analyzed for osteogenic potency after cell culture. The results suggested that ASCs exhibit distinct populations with differing characteristics for osteogenic differentiation: unsorted ASCs stimulated comparable mineralized nodule formation as bone marrow stromal cells (BMSCs) in osteogenic medium; and viral transfection for BMP2 accelerated the formation of mineralized nodules in CD90 and/or CD105 positive ASCs with observation of decrease in CD105 expression after 14 days. Future studies assessing different immunophenotypes of ASCs should be undertaken to develop cell-based tissue engineering.

PMID: 25302065

 

Supplement:

Stem cells have significant potential in multidisciplinary regenerative medicine. However, because many of difficulties, such as establishment of ethic and safety, and irreproducibility still remain before clinical use of stem cells, it is necessary, partially urgent, to pursue ideal stem cells so that they can then be differentiated along multiple cell lineage pathways in a regulated and reproducible manner, transplanted safely and effectively to an autologous or allogeneic host, and manufactured in accordance with current good manufacturing practice guidelines (1). Our research group comprises oral implantologists and often encounter problems that bony recovery is not accomplished enough, and is, indeed, urgent for oral implant prostheses with less traumatic invasive procedures. Therefore, simple, safe and promising strategies for bone healing are definitely required. A series of adipose-derived stem cells (ASCs), which are easily extracted from the buccal fat pad during the implant surgical operation, are strong candidate as a stem cell resource, compared with bone marrow stem cells; because they are not only to meet the stated requirement but also to have been reported that they contain approximately 500 times as dense somatic stem cells as bone marrow.

Figure 1

Figure 1: DNA microarray for wnt-related or osteogenesis-related genes expressed in sorted ASCs.

 

Adipose tissue contains adipocytes, endothelial cells, fibroblasts, immune cells, vascular smooth muscle cells, and ASCs. ASCs are collected in the pelleted stromal vascular fraction (SVF) by centrifugation and resemble mesenchymal stem cells (MSCs) owing to their general differentiation potential and plasticity (2, 3). Furthermore, ASCs display cell surface markers such as the CD family like bone marrow-derived mesenchymal stem cells (BMSCs) representative of somatic stem cells. In this experiment, we brought CD90 and CD105 into focus. Although, CD90 and CD105 can be used to identify stem cells from total SVFs and adipocytes likely to form osteoblastic or chondroblastic progenies (4), only a small number of studies have examined the importance of CDs for stem cell growth and differentiation (5). At the beginning of the study, FACS was performed to separate the ASCs for CD90 and/or CD105, and then DNA microarray profiled genes of interest in each sorted population (Figure 1).

Figure 2

Figure 2: Adenoviral transfection of AdenoX-bmp2 into HEK293 cells (2.5 × 105 cells/500 µL in each well of 24-well plate). The transfected cells were visualized with red fluorescent protein.

 

In this study, murine-derived ASCs were sorted for the expression of the surface protein molecules CD90 and CD105 using FACS. This is the first study to examine the osteogenic potential of the sorted populations with or without bone morphogenetic protein-2 (BMP2) adenoviral transfection (Figure 2) and identified distinct populations of ASCs with differing characteristics for osteogenic differentiation.

Conclusively, the osteogenic potency differed among the populations. Although ASCs and BMSCs represented a similar efficacy, BMP2 significantly increased the formation of mineralized nodules especially in CD90-positive populations with observation of decreased CD105 expression.

 

References

(1)          Gimble JM, Katz AJ, Bunnell BA. Adipose-derived stem cells for regenerative medicine Circ Res 2007; 100: 1249-1260.

(2)          Rodbell M Metabolism of isolated fat cells. II. The similar effects of phospholipase C (Clostridium perfringens alpha toxin) and of insulin on glucose and amino acid metabolism. J Biol Chem 1966; 241: 130-139.

(3)          Rodbell M The metabolism of isolated fat cells. IV. Regulation of release of protein by lipolytic hormones and insulin. J Biol Chem 1966; 241: 3909-3917.

(4)          Mitterberger MC, Lechner S, Mattesich M, et al. DLK1(PREF1) is a negative regulator of adipogenesis in CD105(+)/CD90(+)/CD34(+)/CD31(-)/FABP4(-) adipose-derived stromal cells from subcutaneous abdominal fat pats of adult women. Stem Cell Res 2012; 9: 35-48.

(5)          Huang SJ, Fu RH, Shyu WC, et al. Adipose-derived stem cells: isolation, characterization, and differentiation potential. Cell Transplant 2013; 22: 701-709.

 

 

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