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Tissue Engineering of Ureteral Grafts: Preparation of Biocompatible Crosslinked Ureteral Scaffolds of Porcine Origin.

Koch H, Hammer N, Ossmann S, Schierle K, Sack U, Hofmann J, Wecks M, Boldt A - Front Bioeng Biotechnol (2015)

Bottom Line: After decellularization, scaffold morphology and composition of ECM were maintained, all cellular components were removed, DNA destroyed and strongly reduced.In vitro: GP and CDI scaffolds revealed a higher number of ingrown 3T3 and SMC cells as compared to untreated scaffolds.TIMP1 was below the detection limit.

View Article: PubMed Central - PubMed

Affiliation: Translational Centre for Regenerative Medicine (TRM), University of Leipzig , Leipzig , Germany.

ABSTRACT
The surgical reconstruction of ureteric defects is often associated with post-operative complications and requires additional medical care. Decellularized ureters originating from porcine donors could represent an alternative therapy. Our aim was to investigate the possibility of manufacturing decellularized ureters, the characteristics of the extracellular matrix (ECM) and the biocompatibility of these grafts in vitro/in vivo after treatment with different crosslinking agents. To achieve these goals, native ureters were obtained from pigs and were decellularized. The success of decellularization and the ECM composition were characterized by (immuno)histological staining methods and a DNA-assay. In vitro: scaffolds were crosslinked either with carbodiimide (CDI), genipin (GP), glutaraldehyde, left chemically untreated or were lyophilized. Scaffolds in each group were reseeded with Caco2, LS48, 3T3 cells, or native rat smooth muscle cells (SMC). After 2 weeks, the number of ingrown cells was quantified. In vivo: crosslinked scaffolds were implanted subcutaneously into rats and the type of infiltrating cells were determined after 1, 9, and 30 days. After decellularization, scaffold morphology and composition of ECM were maintained, all cellular components were removed, DNA destroyed and strongly reduced. In vitro: GP and CDI scaffolds revealed a higher number of ingrown 3T3 and SMC cells as compared to untreated scaffolds. In vivo: at day 30, implants were predominantly infiltrated by fibroblasts and M2 anti-inflammatory macrophages. A maximum of MMP3 was observed in the CDI group at day 30. TIMP1 was below the detection limit. In this study, we demonstrated the potential of decellularization to create biocompatible porcine ureteric grafts, whereas a CDI-crosslink may facilitate the remodeling process. The use of decellularized ureteric grafts may represent a novel therapeutic method in reconstruction of ureteric defects.

No MeSH data available.


Related in: MedlinePlus

Histological analysis of HE-stained subcutaneous implanted untreated (A,C,E) and CDI crosslinked scaffolds (B,D,F) explanted at days 1, 9, and 30 post-operative. At day 1 post implantation, cellular infiltration with granulocytes, fibroblasts, and macrophages could be observed in untreated scaffolds (A), whereas in CDI crosslinked scaffolds (B) only a cellular layer was detectable at the periphery. At day 9 post implantation, a notable increase of infiltrating cells into untreated scaffolds (C) was detected as a sign of encapsulation. CDI crosslinked (D) scaffolds showed a mild cellular infiltration with granulocytes, fibroblasts, and macrophages. At day 30 post implantation, in contrast to the untreated scaffold group (E), CDI crosslinked scaffolds (F) showed only a marginal cellular infiltration by granulocytes, fibroblasts, and macrophages. Untreated scaffolds were completely infiltrated by cells and largely degraded. Detailed cellular analysis is displayed in Figure 8. Bar = 100 μm.
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Figure 7: Histological analysis of HE-stained subcutaneous implanted untreated (A,C,E) and CDI crosslinked scaffolds (B,D,F) explanted at days 1, 9, and 30 post-operative. At day 1 post implantation, cellular infiltration with granulocytes, fibroblasts, and macrophages could be observed in untreated scaffolds (A), whereas in CDI crosslinked scaffolds (B) only a cellular layer was detectable at the periphery. At day 9 post implantation, a notable increase of infiltrating cells into untreated scaffolds (C) was detected as a sign of encapsulation. CDI crosslinked (D) scaffolds showed a mild cellular infiltration with granulocytes, fibroblasts, and macrophages. At day 30 post implantation, in contrast to the untreated scaffold group (E), CDI crosslinked scaffolds (F) showed only a marginal cellular infiltration by granulocytes, fibroblasts, and macrophages. Untreated scaffolds were completely infiltrated by cells and largely degraded. Detailed cellular analysis is displayed in Figure 8. Bar = 100 μm.

Mentions: The cell infiltration in untreated (Figures 7A,C,E) and crosslinked scaffolds (Figures 7B,D,F) was analyzed histologically after explantation at days 1, 9, and 30 post implantation. At day 1 after implantation, a cellular infiltration directed from the periphery to the center of the respective tissue was observed in untreated scaffolds (Figure 7A). After crosslinking with CDI (Figure 7B), a cellular layer was detectable at the periphery and only a few number of cells infiltrated the central parts of the scaffolds. At day 9 post implantation, a decrease of cell infiltration was observed in crosslinked scaffolds (Figure 7D) compared to that in untreated scaffolds (Figure 7C). At day 30 post implantation, the untreated implants were largely degraded and completely infiltrated with cells (Figure 7E). In the CDI crosslinked group only an immaterial cellular infiltration could be detected (Figure 7F). In all crosslinked groups (GP, GA, CDI, BP), the cell infiltration was lower compared to the untreated scaffold group but without significant differences among the groups.


Tissue Engineering of Ureteral Grafts: Preparation of Biocompatible Crosslinked Ureteral Scaffolds of Porcine Origin.

Koch H, Hammer N, Ossmann S, Schierle K, Sack U, Hofmann J, Wecks M, Boldt A - Front Bioeng Biotechnol (2015)

Histological analysis of HE-stained subcutaneous implanted untreated (A,C,E) and CDI crosslinked scaffolds (B,D,F) explanted at days 1, 9, and 30 post-operative. At day 1 post implantation, cellular infiltration with granulocytes, fibroblasts, and macrophages could be observed in untreated scaffolds (A), whereas in CDI crosslinked scaffolds (B) only a cellular layer was detectable at the periphery. At day 9 post implantation, a notable increase of infiltrating cells into untreated scaffolds (C) was detected as a sign of encapsulation. CDI crosslinked (D) scaffolds showed a mild cellular infiltration with granulocytes, fibroblasts, and macrophages. At day 30 post implantation, in contrast to the untreated scaffold group (E), CDI crosslinked scaffolds (F) showed only a marginal cellular infiltration by granulocytes, fibroblasts, and macrophages. Untreated scaffolds were completely infiltrated by cells and largely degraded. Detailed cellular analysis is displayed in Figure 8. Bar = 100 μm.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4477215&req=5

Figure 7: Histological analysis of HE-stained subcutaneous implanted untreated (A,C,E) and CDI crosslinked scaffolds (B,D,F) explanted at days 1, 9, and 30 post-operative. At day 1 post implantation, cellular infiltration with granulocytes, fibroblasts, and macrophages could be observed in untreated scaffolds (A), whereas in CDI crosslinked scaffolds (B) only a cellular layer was detectable at the periphery. At day 9 post implantation, a notable increase of infiltrating cells into untreated scaffolds (C) was detected as a sign of encapsulation. CDI crosslinked (D) scaffolds showed a mild cellular infiltration with granulocytes, fibroblasts, and macrophages. At day 30 post implantation, in contrast to the untreated scaffold group (E), CDI crosslinked scaffolds (F) showed only a marginal cellular infiltration by granulocytes, fibroblasts, and macrophages. Untreated scaffolds were completely infiltrated by cells and largely degraded. Detailed cellular analysis is displayed in Figure 8. Bar = 100 μm.
Mentions: The cell infiltration in untreated (Figures 7A,C,E) and crosslinked scaffolds (Figures 7B,D,F) was analyzed histologically after explantation at days 1, 9, and 30 post implantation. At day 1 after implantation, a cellular infiltration directed from the periphery to the center of the respective tissue was observed in untreated scaffolds (Figure 7A). After crosslinking with CDI (Figure 7B), a cellular layer was detectable at the periphery and only a few number of cells infiltrated the central parts of the scaffolds. At day 9 post implantation, a decrease of cell infiltration was observed in crosslinked scaffolds (Figure 7D) compared to that in untreated scaffolds (Figure 7C). At day 30 post implantation, the untreated implants were largely degraded and completely infiltrated with cells (Figure 7E). In the CDI crosslinked group only an immaterial cellular infiltration could be detected (Figure 7F). In all crosslinked groups (GP, GA, CDI, BP), the cell infiltration was lower compared to the untreated scaffold group but without significant differences among the groups.

Bottom Line: After decellularization, scaffold morphology and composition of ECM were maintained, all cellular components were removed, DNA destroyed and strongly reduced.In vitro: GP and CDI scaffolds revealed a higher number of ingrown 3T3 and SMC cells as compared to untreated scaffolds.TIMP1 was below the detection limit.

View Article: PubMed Central - PubMed

Affiliation: Translational Centre for Regenerative Medicine (TRM), University of Leipzig , Leipzig , Germany.

ABSTRACT
The surgical reconstruction of ureteric defects is often associated with post-operative complications and requires additional medical care. Decellularized ureters originating from porcine donors could represent an alternative therapy. Our aim was to investigate the possibility of manufacturing decellularized ureters, the characteristics of the extracellular matrix (ECM) and the biocompatibility of these grafts in vitro/in vivo after treatment with different crosslinking agents. To achieve these goals, native ureters were obtained from pigs and were decellularized. The success of decellularization and the ECM composition were characterized by (immuno)histological staining methods and a DNA-assay. In vitro: scaffolds were crosslinked either with carbodiimide (CDI), genipin (GP), glutaraldehyde, left chemically untreated or were lyophilized. Scaffolds in each group were reseeded with Caco2, LS48, 3T3 cells, or native rat smooth muscle cells (SMC). After 2 weeks, the number of ingrown cells was quantified. In vivo: crosslinked scaffolds were implanted subcutaneously into rats and the type of infiltrating cells were determined after 1, 9, and 30 days. After decellularization, scaffold morphology and composition of ECM were maintained, all cellular components were removed, DNA destroyed and strongly reduced. In vitro: GP and CDI scaffolds revealed a higher number of ingrown 3T3 and SMC cells as compared to untreated scaffolds. In vivo: at day 30, implants were predominantly infiltrated by fibroblasts and M2 anti-inflammatory macrophages. A maximum of MMP3 was observed in the CDI group at day 30. TIMP1 was below the detection limit. In this study, we demonstrated the potential of decellularization to create biocompatible porcine ureteric grafts, whereas a CDI-crosslink may facilitate the remodeling process. The use of decellularized ureteric grafts may represent a novel therapeutic method in reconstruction of ureteric defects.

No MeSH data available.


Related in: MedlinePlus