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Urethral reconstruction with tissue-engineered human amniotic scaffold in rabbit urethral injury models.

Wang F, Liu T, Yang L, Zhang G, Liu H, Yi X, Yang X, Lin TY, Qin W, Yuan J - Med. Sci. Monit. (2014)

Bottom Line: After the successful acquisition of dHAS from AM, cell-seeded dHAS were prepared and characterized.Immune responses were compared by histological evaluation and CD4 cell and CD8 cell infiltrations.Histopathological analysis revealed mild immune response in cell-seeded dHAS group (P<0.05).

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, Xijing Hospital, Fourth Military Medical University, Xi'an, China (mainland).

ABSTRACT

Background: Mitigating urethral injury remains a great challenge for urologists due to lack of ideal biomaterials for urethroplasty. The application of amniotic membranes (AM) over other synthetic materials make it a better potential source for urethral reconstruction. We separated the basement layer of AM to obtain denuded human amniotic scaffold (dHAS) and then inoculated primary rabbit urethral epithelial cells on the surface of dHAS to define whether this strategy minimize potential rejection and maximize the biocompatibility of human AM.

Material/methods: After the successful acquisition of dHAS from AM, cell-seeded dHAS were prepared and characterized. Both cell-seeded dHAS and acellular dHAS were subcutaneously implanted. Immune responses were compared by histological evaluation and CD4 cell and CD8 cell infiltrations. Then they were applied as urethroplastic materials in the rabbit models of urethral injury to fully explore the feasibility and efficacy of tissue-engineered dHAS xenografts in urethral substitution application.

Results: Mild inflammatory infiltration was observed in cell-seeded dHAS grafts, as revealed by fewer accumulations of CD4 cells and CD8 cells (or neutrophils or other immune cells). Urethral defects of rabbits in the urethroplastic group with dHAS implantation (n=6) were completely resolved in one month, while there were one infection and one fistula in the control group with acellular dHAS patches (n=6). Histopathological analysis revealed mild immune response in cell-seeded dHAS group (P<0.05).

Conclusions: Tissue-engineered dHAS minimize potential rejection and maximize the biocompatibility of AM, which makes it a potential ideal xenograft for urethral reconstruction.

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Related in: MedlinePlus

(A–F) Characterizations of denuded human amniotic scaffold (dHAS). (A) Scanning electron micrograph of dHAS showed a continuous flat layer of smooth basement membrane; (B) Hematoxylin-eosin (HE) staining of dHAS confirmed a complete removal of the epithelial layer of amniotic membrane (AM); (C) DAPI staining revealed the absence of epithelial cells; (D) immunofluorescence of Collagen I expression in dHAS; (E) Collagen III; (F) Collagen IV. (G–I) Characterizations of denuded human amniotic scaffold (dHAS). (G) Fibronectin; (H) VEGF; (I) negative control group incubated with nonspecific IgG (magnification at ×400).
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f1-medscimonit-20-2430: (A–F) Characterizations of denuded human amniotic scaffold (dHAS). (A) Scanning electron micrograph of dHAS showed a continuous flat layer of smooth basement membrane; (B) Hematoxylin-eosin (HE) staining of dHAS confirmed a complete removal of the epithelial layer of amniotic membrane (AM); (C) DAPI staining revealed the absence of epithelial cells; (D) immunofluorescence of Collagen I expression in dHAS; (E) Collagen III; (F) Collagen IV. (G–I) Characterizations of denuded human amniotic scaffold (dHAS). (G) Fibronectin; (H) VEGF; (I) negative control group incubated with nonspecific IgG (magnification at ×400).

Mentions: Trypsinization with mixed digestive solution successfully removed the epithelial layer of the amniotic membrane, as revealed by SEM and HE staining (Figure 1A, 1B). Nuclei staining using DAPI revealed the absence of epithelial cells (Figure 1C). The expression patterns of extracellular matrix molecules and growth factor in dHAS were characterized by immunofluorescence imaging (Figure 1C–1I). Ubiquitous expressions of collagen (types I, III, and IV), fibronectin, and vascular endothelial growth factor (VEGF) were demonstrated, although fibronectin and VEGF expressions were lower compared to collagen expression. No positive staining was found under the microscopic field in negative controls.


Urethral reconstruction with tissue-engineered human amniotic scaffold in rabbit urethral injury models.

Wang F, Liu T, Yang L, Zhang G, Liu H, Yi X, Yang X, Lin TY, Qin W, Yuan J - Med. Sci. Monit. (2014)

(A–F) Characterizations of denuded human amniotic scaffold (dHAS). (A) Scanning electron micrograph of dHAS showed a continuous flat layer of smooth basement membrane; (B) Hematoxylin-eosin (HE) staining of dHAS confirmed a complete removal of the epithelial layer of amniotic membrane (AM); (C) DAPI staining revealed the absence of epithelial cells; (D) immunofluorescence of Collagen I expression in dHAS; (E) Collagen III; (F) Collagen IV. (G–I) Characterizations of denuded human amniotic scaffold (dHAS). (G) Fibronectin; (H) VEGF; (I) negative control group incubated with nonspecific IgG (magnification at ×400).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4257484&req=5

f1-medscimonit-20-2430: (A–F) Characterizations of denuded human amniotic scaffold (dHAS). (A) Scanning electron micrograph of dHAS showed a continuous flat layer of smooth basement membrane; (B) Hematoxylin-eosin (HE) staining of dHAS confirmed a complete removal of the epithelial layer of amniotic membrane (AM); (C) DAPI staining revealed the absence of epithelial cells; (D) immunofluorescence of Collagen I expression in dHAS; (E) Collagen III; (F) Collagen IV. (G–I) Characterizations of denuded human amniotic scaffold (dHAS). (G) Fibronectin; (H) VEGF; (I) negative control group incubated with nonspecific IgG (magnification at ×400).
Mentions: Trypsinization with mixed digestive solution successfully removed the epithelial layer of the amniotic membrane, as revealed by SEM and HE staining (Figure 1A, 1B). Nuclei staining using DAPI revealed the absence of epithelial cells (Figure 1C). The expression patterns of extracellular matrix molecules and growth factor in dHAS were characterized by immunofluorescence imaging (Figure 1C–1I). Ubiquitous expressions of collagen (types I, III, and IV), fibronectin, and vascular endothelial growth factor (VEGF) were demonstrated, although fibronectin and VEGF expressions were lower compared to collagen expression. No positive staining was found under the microscopic field in negative controls.

Bottom Line: After the successful acquisition of dHAS from AM, cell-seeded dHAS were prepared and characterized.Immune responses were compared by histological evaluation and CD4 cell and CD8 cell infiltrations.Histopathological analysis revealed mild immune response in cell-seeded dHAS group (P<0.05).

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, Xijing Hospital, Fourth Military Medical University, Xi'an, China (mainland).

ABSTRACT

Background: Mitigating urethral injury remains a great challenge for urologists due to lack of ideal biomaterials for urethroplasty. The application of amniotic membranes (AM) over other synthetic materials make it a better potential source for urethral reconstruction. We separated the basement layer of AM to obtain denuded human amniotic scaffold (dHAS) and then inoculated primary rabbit urethral epithelial cells on the surface of dHAS to define whether this strategy minimize potential rejection and maximize the biocompatibility of human AM.

Material/methods: After the successful acquisition of dHAS from AM, cell-seeded dHAS were prepared and characterized. Both cell-seeded dHAS and acellular dHAS were subcutaneously implanted. Immune responses were compared by histological evaluation and CD4 cell and CD8 cell infiltrations. Then they were applied as urethroplastic materials in the rabbit models of urethral injury to fully explore the feasibility and efficacy of tissue-engineered dHAS xenografts in urethral substitution application.

Results: Mild inflammatory infiltration was observed in cell-seeded dHAS grafts, as revealed by fewer accumulations of CD4 cells and CD8 cells (or neutrophils or other immune cells). Urethral defects of rabbits in the urethroplastic group with dHAS implantation (n=6) were completely resolved in one month, while there were one infection and one fistula in the control group with acellular dHAS patches (n=6). Histopathological analysis revealed mild immune response in cell-seeded dHAS group (P<0.05).

Conclusions: Tissue-engineered dHAS minimize potential rejection and maximize the biocompatibility of AM, which makes it a potential ideal xenograft for urethral reconstruction.

Show MeSH
Related in: MedlinePlus