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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 the degree of scaffold infiltration by giant cells (A), granulocytes (B), capillaries (C), collagen fibers (D), lymphocytes (E), and fibroblasts (F) at days 1, 9, and 30 post implantation. In all implants, an increase of infiltrating giant cells, lymphocytes, fibroblasts, capillaries, and collagen fibers could be detected over time. Furthermore, a decrease of infiltration by granulocytes could be observed in all implants at day 30 compared to day 1 post implantation. A blinded pathologist evaluated the presence of cells (granulocytes, lymphocytes, giant cells, and fibroblasts), capillaries and new collagen fibers and generated a semiquantitative score in repeat determination (0 = negative, 1 = mild, 2 = moderate, 3 = serve; n = 6/group and day; five microscopic fields per slide and rat; each 5 mm2). Untreated, untreated decellular ureteral tissue; BP, bovine pericardium (St. Jude, USA); GP, genipin; GA, glutaraldehyde; CDI, carbodiimide. **P < 0.01 vs. untreated decellular ureteral tissue.
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Figure 8: Histological analysis of the degree of scaffold infiltration by giant cells (A), granulocytes (B), capillaries (C), collagen fibers (D), lymphocytes (E), and fibroblasts (F) at days 1, 9, and 30 post implantation. In all implants, an increase of infiltrating giant cells, lymphocytes, fibroblasts, capillaries, and collagen fibers could be detected over time. Furthermore, a decrease of infiltration by granulocytes could be observed in all implants at day 30 compared to day 1 post implantation. A blinded pathologist evaluated the presence of cells (granulocytes, lymphocytes, giant cells, and fibroblasts), capillaries and new collagen fibers and generated a semiquantitative score in repeat determination (0 = negative, 1 = mild, 2 = moderate, 3 = serve; n = 6/group and day; five microscopic fields per slide and rat; each 5 mm2). Untreated, untreated decellular ureteral tissue; BP, bovine pericardium (St. Jude, USA); GP, genipin; GA, glutaraldehyde; CDI, carbodiimide. **P < 0.01 vs. untreated decellular ureteral tissue.

Mentions: 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.


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 the degree of scaffold infiltration by giant cells (A), granulocytes (B), capillaries (C), collagen fibers (D), lymphocytes (E), and fibroblasts (F) at days 1, 9, and 30 post implantation. In all implants, an increase of infiltrating giant cells, lymphocytes, fibroblasts, capillaries, and collagen fibers could be detected over time. Furthermore, a decrease of infiltration by granulocytes could be observed in all implants at day 30 compared to day 1 post implantation. A blinded pathologist evaluated the presence of cells (granulocytes, lymphocytes, giant cells, and fibroblasts), capillaries and new collagen fibers and generated a semiquantitative score in repeat determination (0 = negative, 1 = mild, 2 = moderate, 3 = serve; n = 6/group and day; five microscopic fields per slide and rat; each 5 mm2). Untreated, untreated decellular ureteral tissue; BP, bovine pericardium (St. Jude, USA); GP, genipin; GA, glutaraldehyde; CDI, carbodiimide. **P < 0.01 vs. untreated decellular ureteral tissue.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4477215&req=5

Figure 8: Histological analysis of the degree of scaffold infiltration by giant cells (A), granulocytes (B), capillaries (C), collagen fibers (D), lymphocytes (E), and fibroblasts (F) at days 1, 9, and 30 post implantation. In all implants, an increase of infiltrating giant cells, lymphocytes, fibroblasts, capillaries, and collagen fibers could be detected over time. Furthermore, a decrease of infiltration by granulocytes could be observed in all implants at day 30 compared to day 1 post implantation. A blinded pathologist evaluated the presence of cells (granulocytes, lymphocytes, giant cells, and fibroblasts), capillaries and new collagen fibers and generated a semiquantitative score in repeat determination (0 = negative, 1 = mild, 2 = moderate, 3 = serve; n = 6/group and day; five microscopic fields per slide and rat; each 5 mm2). Untreated, untreated decellular ureteral tissue; BP, bovine pericardium (St. Jude, USA); GP, genipin; GA, glutaraldehyde; CDI, carbodiimide. **P < 0.01 vs. untreated decellular ureteral tissue.
Mentions: 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.

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