Fixation of porous calcium phosphate with expanded bone marrow cells using an autologous plasma clot

Abstract

Multipotent human mesenchymal stromal cells (hMSCs) are currently the most promising cell type for regenerative medicine and tissue engineering. For clinical applications with special focus on fracture repair a method to deliver in vitro-expanded bone marrow cells to the fracture site is presented. Autologous blood plasma clotting was used to fix expanded cells and synthetic tricalcium phosphate particles. A jelly-like but solid composite matrix was obtained which could easily be handled by the surgeon. The majority of embedded cells remained viable after the plasma gelation process, as was determined by fluorescence microscopy. Experimental cell culture over a period of 15 days showed proliferating mesenchymal stromal cells within the plasma clot.Multipotente mesenchymale Stromazellen gelten als viel versprechender Zelltyp für regenerative Medizin und Tissue Engineering. Im Rahmen klinischer Anwendungen mit Fokus auf die Frakturheilung wird eine Methode zur Versorgung mit in vitro-expandierten Knochenmarkszellen in Frakturnähe vorgestellt. Zur Fixierung der expandierten Zellen zusammen mit synthetischer Tricalciumphosphat-Granula wurde autologes Blutplasma verwendet und so eine gellee-artige aber feste Komposit-Matrix hergestellt, die vom Chirurgen einfach zu handhaben war. Wie mittels Fluoreszenzmikroskopie bestimmt wurde, blieb die Mehrzahl der eingebetteten Zellen nach dem Gelierungsprozess vital. Eine anschließende experimentelle Zellkultur für weitere 15 Tage zeigte im Plasmakoagulat proliferierende mesenchymale Stromazellen.

Osteogenic differentiation of human mesenchymal stromal cells is promoted by a leukocytes containing fibrin matrix

“…It is likely that the presence of different platelet/leukocyte number at the time of gelation profoundly influences the plasma clot structure. We have shown that also nonviable cells are detectable at distinct and localized areas after the clotting process within a plasma matrix which obviously resulted from cell squeezing due to the formation of fibrin fiber bundles [3]. Clots formed in the presence of low thrombin concentrations are composed of thick fibrin fibers, while clots formed in the presence of high thrombin concentrations are composed of thin fibers [25].…”

“…On the other hand, there are few drawbacks with the use of plasma clots which consist of completely autologous material. In contrast to commercially available fibrin matrices or sealants which are adjustable in matrix strength by using different concentrations of fibrinogen and thrombin [24], an autologous plasma clot is more heterogeneous in its microstructure [3]. Here, varying factors will influence the dynamic process of the clot formation which includes the content of platelets and/or leukocytes.…”

“…The major advantage of a plasma clot is the lack of allogenic or xenogenic factors during clot preparation which are present and necessary in commercially available fibrin sealant systems. The feasibility of such a plasma clot system under clinical conditions has been shown by our group [3]. In addition, we have also shown that isolated and subsequently lipopolysaccharide (LPS)-activated peripheral blood mononuclear leukocytes (PBMC) released soluble factors or a combination of factors which are able to induce osteogenic differentiation of MSC in vitro [6].…”

“…The plasma clot exerts a three-dimensional microporous structure and excellent biocompatibility and allows diffusion of regulatory factors [3,4]. Furthermore, a plasma clot system will allow proliferation and probably differentiation of embedded MSC [3,5].…”

“…Furthermore, a plasma clot system will allow proliferation and probably differentiation of embedded MSC [3,5]. The major advantage of a plasma clot is the lack of allogenic or xenogenic factors during clot preparation which are present and necessary in commercially available fibrin sealant systems.…”