Naslov
Tracking Osteocyte Lineage Plasticity In Vitro

Autori
Pejda, Slavica ; Kizivat, Tomislav ; Fatahi, Mohammad ; Igwe, John ; Kalajzić, Ivo

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
The American Society for Bone and Mineral Research 2010. Annual Meeting Abstract Book / - Hoboken (NJ) : John Wiley & Sons, 2010, S113-S113

Skup
The American Society for Bone and Mineral Research 2010. Annual Meeting

Mjesto i datum
Toronto, Kanada, 15.10.2010. - 19.10.2010

Vrsta sudjelovanja
Poster

Vrsta recenzije
Međunarodna recenzija

Ključne riječi
bone; osteocyte; transgenical mice; cell plasticity; bone chip; de-differentiation; osteoblasts

Sažetak
Presently, there is no evidence that mature osteoblast and osteocytes are able to dedifferentiate. However, dedifferentiation of these cells could provide an additional source of cells for skeletal reconstruction/repair. The use of visual transgenes, such as GFP, has been instrumental in identifying cells at different stages of maturation. However, markers that are under the control of specific promoters that are no longer expressed after the cell reaches a point when promoter activity ceases, or when a cell transitions to an alternative differentiation state cannot be used to track cell histories. It is then necessary to to use a tool like the Cre/loxP system for lineage tracing. In this study we present evidence that preosteocytes/osteocytes can undergo dedifferentiation, accompanied by dramatic changes in gene expression. We have utilized previously developed transgenic mice in which Dmp1 promoter had been used with GFP as a visual marker of current stage of differentiation (DMP-GFP) or to drive Cre recombinase and turn on an historical marker in osteocytes and odontoblasts (DMP-Cre). Primary bone chip outgrowth cell (BCOC) cultures were prepared after surface osteoblasts have been removed using enzymatic digestion. When we utilized a DMP-GFP transgene, outgrowth cells were GFP negative by epifluorescence and by immunofluorescent staining using GFP antibody. Using DMP-Cre mice, floxed GFPreporter+ cells emerged from bone chips after 48-72 hours, where GFP expression iindicates their previous osteocytic phenotype. In addition, a large proportion of BCOC’s began to express a-SMA-GFP. Preliminary results obtained by microarray analysis of BCOC isolated 7 days after plating and compared to the RNA of marrow stromal cells showed large increases in markers characteristic of mature osteoblast/osteocytes, including Bglap, Ibsp, Phex, Osterix, BMP8a, and Pthr1. Expanded BCOC’s in vitro were transplanted by intra-bone marrow cavity injection. Four weeks after transplantation, GFP-labeled donor cells were detected as numerous osteocytes deeply embedded in newly formed bone. Our results indicate that under certain conditions cells embedded in the matrix can emerge out and begin to proliferate. This process does not preclude the ability of outgrowth cells to differentiate again into mature osteocytes. Future studies should be aimed at evaluating factors that affect the process of dedifferentiation with the goal of increasing bone formation in vivo and in vitro.

Izvorni jezik
Engleski

Znanstvena područja
Temeljne medicinske znanosti

POVEZANOST RADA