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Dive into the research topics where Carina Orth is active.

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Featured researches published by Carina Orth.


Biomaterials | 2009

Contribution of outgrowth endothelial cells from human peripheral blood on in vivo vascularization of bone tissue engineered constructs based on starch polycaprolactone scaffolds

Sabine Fuchs; Shahram Ghanaati; Carina Orth; Mike Barbeck; Marlen Kolbe; Alexander Hofmann; Markus Eblenkamp; Manuela E. Gomes; Rui L. Reis; Charles James Kirkpatrick

In the present study we assessed the potential of human outgrowth endothelial cells (OEC), a subpopulation within endothelial progenitor cell cultures, to support the vascularization of a complex tissue engineered construct for bone. OEC cultured on starch polycaprolactone fiber meshes (SPCL) in monoculture retained their endothelial functionality and responded to angiogenic stimulation by VEGF (vascular endothelial growth factor) in fibrin gel-assays in vitro. Co-culture of OEC with human primary osteoblasts (pOB) on SPCL, induced an angiogenic activation of OEC towards microvessel-like structures achieved without additional supplementation with angiogenic growth factors. Effects of co-cultures with pOB on the vascularization process by OEC in vivo were tested by subcutaneous implantation of Matrigel plugs containing both, OEC and pOB, and resulted in OEC-derived blood vessels integrated into the host tissue and anastomosed to the vascular supply. In addition, morphometric analysis of the vascularization process by OEC indicated a better performance of OEC in the co-cultures with primary osteoblasts compared to monocultures of OEC. The contribution of OEC to vascular structures and the beneficial effect of the co-culture with primary human osteoblasts on the vascularization in vivo was additionally proven by subcutaneous implantation of pre-cellularized and pre-cultured SPCL constructs. OEC contributed to the vascular structures, by generating autogenic vessels or by incorporation into chimeric vessels consisting of both, human and mouse endothelial cells. The current data highlight the vasculogenic potential of OEC for bone tissue engineering applications and indicate a beneficial influence of constructs including both osteoblasts and endothelial cells for vascularization strategies.


Biomaterials | 2010

The rapid anastomosis between prevascularized networks on silk fibroin scaffolds generated in vitro with cocultures of human microvascular endothelial and osteoblast cells and the host vasculature

Ronald E. Unger; Shahram Ghanaati; Carina Orth; Anne Sartoris; Mike Barbeck; Sven Halstenberg; Antonella Motta; Claudio Migliaresi; C. James Kirkpatrick

The survival and functioning of a bone biomaterial upon implantation requires a rapidly forming and stably functioning vascularization that connects the implant to the recipient. We have previously shown that human microcapillary endothelial cells (HDMEC) and primary human osteoblast cells (HOS) in coculture on various 3-D bone biomaterial scaffolds rapidly distribute and self-assemble into a morphological structure resembling bone tissue. Endothelial cells form microcapillary-like structures containing a lumen and these were intertwined between the osteoblast cells and the biomaterial. This tissue-like self-assembly occurred in the absence of exogenously added angiogenic stimuli or artificial matrices. The purpose of this study was to determine whether this in vitro pre-formed microvasculature persists and functions in vivo and to determine how the host responds to the cell-containing scaffolds. The scaffolds with cocultures were implanted into immune-deficient mice and compared to scaffolds without cells or with HDMEC alone. Histological evaluation and immunohistochemical staining with human-specific antibodies of materials removed 14 days after implantation demonstrated that the in vitro pre-formed microcapillary structures were present on the silk fibroin scaffolds and showed a perfused lumen that contained red blood cells. This proved anastomosis with the host vasculature. Chimeric vessels in which HDMEC were integrated with the hosts ingrowing (murine) capillaries were also observed. No HDMEC-derived microvessel structures or chimeric vessels were observed on implanted silk fibroin when precultured with HDMEC alone. In addition, there was migration of the host (murine) vasculature into the silk fibroin scaffolds implanted with cocultures, whereas silk fibroin alone or silk fibroin precultured only with HDMEC were nearly devoid of ingrowing host microcapillaries. Therefore, not only do the in vitro pre-formed microcapillaries in a coculture survive and anastomose with the host vasculature to become functioning microcapillaries after implantation, the coculture also stimulates the host capillaries to rapidly grow into the scaffold to vascularize the implanted material. Thus, this coculture-based pre-vascularization of a biomaterial implant may have great potential in the clinical setting to treat large bone defects.


Biomaterials | 2009

Dynamic processes involved in the pre-vascularization of silk fibroin constructs for bone regeneration using outgrowth endothelial cells

Sabine Fuchs; Xin Jiang; Harald Schmidt; Eva Dohle; Shahram Ghanaati; Carina Orth; Alexander Hofmann; Antonella Motta; Claudio Migliaresi; Charles James Kirkpatrick

For successful bone regeneration tissue engineered bone constructs combining both aspects, namely a high osteogenic potential and a rapid connection to the vascular network are needed. In this study we assessed the formation of pre-vascular structures by human outgrowth endothelial cells (OEC) from progenitors in the peripheral blood and the osteogenic differentiation of primary human osteoblasts (pOB) on micrometric silk fibroin scaffolds. The rational was to gain more insight into the dynamic processes involved in the differentiation and functionality of both cell types depending on culture time in vitro. Vascular tube formation by OEC was assessed quantitatively at one and 4 weeks of culture. In parallel, we assessed the temporal changes in cell ratios by flow cytometry and in the marker profiles of endothelial and osteogenic markers by quantitative real-time PCR. In terms of OEC, we observed an increase in tube length, tube area, number of nodes and number of vascular meshes within a culture period of 4 weeks, but a decrease in endothelial markers in real-time PCR. At the same time early osteogenic markers were downregulated, while marker expression associated with progressing mineralized matrix was upregulated in later stages of the culture. In addition, deposition of matrix components, such as collagen type I, known as a pro-angiogenic substrate for endothelial cells, appeared to increase with time indicated by immunohistochemistry. In summary, the study suggests a progressing maturation of the tissue construct with culture time which seems to be not effected by culture conditions mainly designed for outgrowth endothelial cells.


Biomaterials | 2009

Dynamic in vivo biocompatibility of angiogenic peptide amphiphile nanofibers

Shahram Ghanaati; Matthew J. Webber; Ronald E. Unger; Carina Orth; James F. Hulvat; Sarah E. Kiehna; Mike Barbeck; Angela Rasic; Samuel I. Stupp; C. James Kirkpatrick

Biomaterials that promote angiogenesis have great potential in regenerative medicine for rapid revascularization of damaged tissue, survival of transplanted cells, and healing of chronic wounds. Supramolecular nanofibers formed by self-assembly of a heparin-binding peptide amphiphile and heparan sulfate-like glycosaminoglycans were evaluated here using a dorsal skinfold chamber model to dynamically monitor the interaction between the nanofiber gel and the microcirculation, representing a novel application of this model. We paired this model with a conventional subcutaneous implantation model for static histological assessment of the interactions between the gel and host tissue. In the static analysis, the heparan sulfate-containing nanofiber gels were found to persist in the tissue for up to 30 days and revealed excellent biocompatibility. Strikingly, as the nanofiber gel biodegraded, we observed the formation of a de novo vascularized connective tissue. In the dynamic experiments using the dorsal skinfold chamber, the material again demonstrated good biocompatibility, with minimal dilation of the microcirculation and only a few adherent leukocytes, monitored through intravital fluorescence microscopy. The new application of the dorsal skinfold model corroborated our findings from the traditional static histology, demonstrating the potential use of this technique to dynamically evaluate the biocompatibility of materials. The observed biocompatibility and development of new vascularized tissue using both techniques demonstrates the potential of these angiogenesis-promoting materials for a host of regenerative strategies.


Acta Biomaterialia | 2010

Influence of β-tricalcium phosphate granule size and morphology on tissue reaction in vivo.

Shahram Ghanaati; Mike Barbeck; Carina Orth; Ines Willershausen; Benjamin W. Thimm; Christiane Hoffmann; Angela Rasic; Robert Sader; Ronald E. Unger; Fabian Peters; C. James Kirkpatrick

In this study the tissue reaction to five different β-tricalcium phosphate (β-TCP)-based bone substitute materials differing only in size, shape and porosity was analyzed over 60 days, at 3, 10, 15, 30 and 60 days after implantation. Using the subcutaneous implantation model in Wistar rats both the inflammatory response within the implantation bed and the resulting vascularization of the biomaterials were qualitatively and quantitatively assessed by means of standard and special histological staining methods. The data from this study showed that all investigated β-TCP bone substitutes induced the formation of multinucleated giant cells. Changes in size, shape and porosity influenced the integration of the biomaterials within the implantation bed and the formation of tartrate-resistant acid phosphatase (TRAP)-positive and TRAP-negative multinucleated giant cells, as well as the rate of vascularization. While a high porosity generally allowed cell and fiber in-growth within the center of the bone substitute, a lower porosity resulted in a mosaic-like integration of the materials, with the granules serving as place holders. The number of multinucleated giant cells located in the implantation bed positively correlated with the vascularization rate. These data emphasize that all biomaterials investigated were capable of inducing the formation of TRAP-positive multinucleated giant cells as a sign of biomaterial stability. Furthermore, these cells directly influenced vascularization by secretion of vascular endothelial growth factor (VEGF), as well as other chemokines. Based on these findings, the role of multinucleated giant cells in the foreign body reaction to biomaterials might need to be reconsidered. This study demonstrates that variations in the physical properties of a bone substitute material clearly influence the (extent of the) inflammatory reaction and its consequences.


Biomaterials | 2011

Scaffold vascularization in vivo driven by primary human osteoblasts in concert with host inflammatory cells.

Shahram Ghanaati; Ronald E. Unger; Matthew J. Webber; Mike Barbeck; Carina Orth; Jenny A. Kirkpatrick; Patrick Booms; Antonella Motta; Claudio Migliaresi; Robert Sader; C. James Kirkpatrick

Successful cell-based tissue engineering requires a rapid and thorough vascularization in order to ensure long-term implant survival and tissue integration. The vascularization of a scaffold is a complex process, and is modulated by the presence of transplanted cells, exogenous and endogenous signaling proteins, and the host tissue reaction, among other influencing factors. This paper presents evidence for the significance of pre-seeded osteoblasts for the in vivo vascularization of a biodegradable scaffold. Human osteoblasts, cultured on silk fibroin micronets in vitro, migrated throughout the interconnected pores of the scaffold and produced extensive bone matrix. When these constructs were implanted in SCID mice, a rapid and thorough vascularization of the scaffold by the host blood capillaries occurred. This profound response was not seen for the silk fibroin scaffold alone. Moreover, when the pre-cultivation time of human osteoblasts was reduced from 14 days to only 24 h, the significant effect these cells exerted on vascularization rate in vivo was still detectable. From these studies, we conclude that matrix and soluble factors produced by osteoblasts can serve to instruct host endothelial cells to migrate, proliferate, and initiate the process of scaffold vascularization. This finding represents a potential paradigm shift for the field of tissue engineering, especially in bone, as traditional strategies to enhance scaffold vascularization have focused on endovascular cells and regarded osteoblasts primarily as cell targets for mineralization. In addition, the migration of host macrophages and multinucleated giant cells into the scaffold was also found to influence the vascularization of the biomaterial. Therefore, the robust effect on scaffold vascularization seen by pre-culturing with osteoblasts appears to occur in concert with the pro-angiogenic stimuli arising from host immune cells.


Journal of Tissue Engineering and Regenerative Medicine | 2011

Rapid vascularization of starch–poly(caprolactone) in vivo by outgrowth endothelial cells in co-culture with primary osteoblasts

Shahram Ghanaati; Sabine Fuchs; Matthew J. Webber; Carina Orth; Mike Barbeck; Manuela E. Gomes; Rui L. Reis; C. James Kirkpatrick

The successful integration of in vitro‐generated tissues is dependent on adequate vascularization in vivo. Human outgrowth endothelial cells (OECs) isolated from the mononuclear cell fraction of peripheral blood represent a potent population of circulating endothelial progenitors that could provide a cell source for rapid anastomosis and scaffold vascularization. Our previous work with these cells in co‐culture with primary human osteoblasts has demonstrated their potential to form perfused vascular structures within a starch–poly(caprolactone) biomaterial in vivo. In the present study, we demonstrate the ability of OECs to form perfused vascular structures as early as 48 h following subcutaneous implantation of the biomaterial in vivo. The number of OEC‐derived vessels increased throughout the study, an effect that was independent of the OEC donor. This finding of rapid and thorough OEC‐mediated scaffold vascularization demonstrates the great potential for OEC‐based strategies to promote vascularization in tissue engineering. OECs have the potential to contribute to host‐derived scaffold vascularization, and formed vascular structures at a similar density as those arising from the host. Additionally, immunohistochemical evidence demonstrated the close interaction between OECs and the co‐cultured osteoblasts. In addition to the known paracrine activity osteoblasts have in modulating angiogenesis of co‐cultured OECs, we demonstrate the potential of osteoblasts to provide additional structural support for OEC‐derived vessels, perhaps acting in a pericyte‐like role. Copyright


Journal of Tissue Engineering and Regenerative Medicine | 2010

Fine-tuning scaffolds for tissue regeneration: effects of formic acid processing on tissue reaction to silk fibroin

Shahram Ghanaati; Carina Orth; Ronald E. Unger; Mike Barbeck; Matthew J. Webber; Antonella Motta; Claudio Migliaresi; C. James Kirkpatrick

Formic acid (FA) plays a key role in the preparation of silk fibroin (SF) scaffolds from cocoons of Bombyx mori and is used for fibre distribution. In this study, we used a subcutaneous implantation model in Wistar rats to examine SF scaffolds prepared by treating the degummed cocoon with FA for either 30 or 60 min. The tissue reaction and inflammatory response to SF was assessed by qualitative histology at intervals from 3 to 180 days. Additionally, dynamic biomaterial‐induced vascularization and biomaterial degradation were quantified using a technique for analysing an image of the entire implanted biomaterial. Varying the FA treatment time led to different scaffold morphologies and resulted in two distinct peri‐implant tissue reactions. The 30 min‐treated scaffold was integrated into the surrounding tissue beginning at day 3 after implantation and vascularization increased 10‐fold from 15 to 180 days, while the scaffold was continuously degraded throughout the first 90 days. In contrast, the 60 min‐treated SF scaffold appeared as bulk for the first 90 days after implantation, after which a rapid degradation and vascularization process began. After 180 days, the tissue response was similar for both scaffolds, with eventual formation of a well vascularized connective tissue integrating the SF fibres. This study indicates that by modifying the FA treatment time, the tissue reaction to SF scaffolds can be tailored for different tissue‐engineering applications. The tunability and biocompatibility of SF make it an attractive scaffold for exploration in regenerative medicine and clinical tissue engineering. Copyright


Biomedical Materials | 2010

Histological and histomorphometrical analysis of a silica matrix embedded nanocrystalline hydroxyapatite bone substitute using the subcutaneous implantation model in Wistar rats

Shahram Ghanaati; Carina Orth; Mike Barbeck; Ines Willershausen; Benjamin W. Thimm; Patrick Booms; Stefan Stübinger; Constantin A. Landes; Robert Anton Sader; Charles James Kirkpatrick

The clinical suitability of a bone substitute material is determined by the ability to induce a tissue reaction specific to its composition. The aim of this in vivo study was to analyze the tissue reaction to a silica matrix-embedded, nanocrystalline hydroxyapatite bone substitute.The subcutaneous implantation model in Wistar rats was chosen to assess the effect of silica degradation on the vascularization of the biomaterial and its biodegradation within a time period of 6 months. Already at day 10 after implantation, histomorphometrical analysis showed that the vascularization of the implantation bed reached its peak value compared to all other time points. Both vessel density and vascularization significantly decreased until day 90 after implantation. In this time period, the bone substitute underwent a significant degradation initiated by TRAP-positive and TRAP-negative multinucleated giant cells together with macrophages and lymphocytes. Although no specific tissue reaction could be related to the described silica degradation, the biomaterial was close to being fully degraded without a severe inflammatory response. These characteristics are advantageous for bone regeneration and remodeling processes.


Biomedical Materials | 2010

Collagen-embedded hydroxylapatite- beta-tricalcium phosphate-silicon dioxide bone substitute granules assist rapid vascularization and promote cell growth

Shahram Ghanaati; Benjamin W. Thimm; Ronald E. Unger; Carina Orth; Thomas Kohler; Mike Barbeck; Ralph Müller; C. James Kirkpatrick

In the present study we assessed the biocompatibility in vitro and in vivo of a low-temperature sol-gel-manufactured SiO(2)-based bone graft substitute. Human primary osteoblasts and the osteoblastic cell line, MG63, cultured on the SiO(2) biomatrix in monoculture retained their osteoblastic morphology and cellular functionality in vitro. The effect of the biomaterial in vivo and its vascularization potential was tested subcutaneously in Wistar rats and demonstrated both rapid vascularization and good integration within the peri-implant tissue. Scaffold degradation was progressive during the first month after implantation, with tartrate-resistant acid phosphatase-positive macrophages being present and promoting scaffold degradation from an early stage. This manuscript describes successful osteoblastic growth promotion in vitro and a promising biomaterial integration and vasculogenesis in vivo for a possible therapeutic application of this biomatrix in future clinical studies.

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Shahram Ghanaati

Goethe University Frankfurt

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Mike Barbeck

Goethe University Frankfurt

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Robert Sader

Goethe University Frankfurt

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