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Human Mesenchymal Stem Cells Reendothelialize Porcine Heart Valve Scaffolds: Novel Perspectives in Heart Valve Tissue Engineering.

Lanuti P, Serafini F, Pierdomenico L, Simeone P, Bologna G, Ercolino E, Di Silvestre S, Guarnieri S, Canosa C, Impicciatore GG, Chiarini S, Magnacca F, Mariggiò MA, Pandolfi A, Marchisio M, Di Giammarco G, Miscia S - Biores Open Access (2015)

Bottom Line: Here, we were able to successfully decellularize porcine heart valves, which were then recellularized with both differentiated-WJ-MSCs and HUVECs.Cells were able to positively interact with the natural matrix and demonstrated the surface expression of typical endothelial markers.Altogether, these data suggest that the interaction between a biological scaffold and WJ-MSCs allows the regeneration of a morphologically well-structured endothelium, opening new perspectives in the field of HVTE.

View Article: PubMed Central - PubMed

Affiliation: Center for Aging Science (Ce.S.I.), "Università G. d'Annunzio" Foundation , Chieti, Italy . ; Department of Medicine and Aging Science, School of Medicine and Health Science, University "G. d'Annunzio" Chieti-Pescara , Chieti, Italy . ; StemTeCh Group , Chieti, Italy .

ABSTRACT
Heart valve diseases are usually treated by surgical intervention addressed for the replacement of the damaged valve with a biosynthetic or mechanical prosthesis. Although this approach guarantees a good quality of life for patients, it is not free from drawbacks (structural deterioration, nonstructural dysfunction, and reintervention). To overcome these limitations, the heart valve tissue engineering (HVTE) is developing new strategies to synthesize novel types of valve substitutes, by identifying efficient sources of both ideal scaffolds and cells. In particular, a natural matrix, able to interact with cellular components, appears to be a suitable solution. On the other hand, the well-known Wharton's jelly mesenchymal stem cells (WJ-MSCs) plasticity, regenerative abilities, and their immunomodulatory capacities make them highly promising for HVTE applications. In the present study, we investigated the possibility to use porcine valve matrix to regenerate in vitro the valve endothelium by WJ-MSCs differentiated along the endothelial lineage, paralleled with human umbilical vein endothelial cells (HUVECs), used as positive control. Here, we were able to successfully decellularize porcine heart valves, which were then recellularized with both differentiated-WJ-MSCs and HUVECs. Data demonstrated that both cell types were able to reconstitute a cellular monolayer. Cells were able to positively interact with the natural matrix and demonstrated the surface expression of typical endothelial markers. Altogether, these data suggest that the interaction between a biological scaffold and WJ-MSCs allows the regeneration of a morphologically well-structured endothelium, opening new perspectives in the field of HVTE.

No MeSH data available.


Related in: MedlinePlus

Native porcine endothelium observed by fluorescence microscopy and compared with the decellularized porcine scaffold. (A) Both native endothelium (a) and decellularized cusps (c) were stained by DAPI (blue). Autofluorescence of the matrix (green) was observed both on native endothelium (b) and on decellularized cusps (d). (B) Native endothelium was stained by cell membrane labeling PKH26 (red, a) and DAPI (b, blue). A PKH26 and DAPI merge image is also shown (c). Decellularized cusps were stained by cell membrane labeling PKH26 (red, d) and DAPI (e, blue). A PKH26 and DAPI merge image is also shown (f). Images are representative of three separate experiments. Scale bar: 15 μm. DAPI, 4′,6-diamidino-2-phenylindole.
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f1: Native porcine endothelium observed by fluorescence microscopy and compared with the decellularized porcine scaffold. (A) Both native endothelium (a) and decellularized cusps (c) were stained by DAPI (blue). Autofluorescence of the matrix (green) was observed both on native endothelium (b) and on decellularized cusps (d). (B) Native endothelium was stained by cell membrane labeling PKH26 (red, a) and DAPI (b, blue). A PKH26 and DAPI merge image is also shown (c). Decellularized cusps were stained by cell membrane labeling PKH26 (red, d) and DAPI (e, blue). A PKH26 and DAPI merge image is also shown (f). Images are representative of three separate experiments. Scale bar: 15 μm. DAPI, 4′,6-diamidino-2-phenylindole.

Mentions: To decellularize heart cusps, two different detergent solutions, already reported in the literature, were compared: one composed of 1% SDS+0.05% NaN3,4 and another containing 1% Triton X-100.17,27 To prove the efficacy of the decellularization process, detergent-treated valves were observed by optical microscopy. Figure 1A-c evidences the complete suppression of the porcine endothelium by fluorescence microscopy since nuclei, pointed out by DAPI staining (blue) on the native cusps (Fig. 1A-a), disappeared when decellularized cusps were observed (Fig. 1A-c). In the green channel, the absence of cell nuclei (Fig. 1A-d) underlined the autofluorescence of the decellularized matrix; such a green autofluorescence disappeared when porcine valve are covered by the native endothelium (Fig. 1A-b). Data were confirmed by multiphoton microscopy analyses, showing the absence of the cellular component on the scaffold, after staining the decellularized valves with PKH26 (red) and DAPI (blue) (Fig. 1B-d–f); the native porcine endothelium was used as positive control for nuclei and membrane staining (Fig. 1B-a–c). Of note, the absence of PKH26-positive elements on decellularized cusps revealed that nor cells neither cellular fragments (which may induce immune reactions) were evidenced.


Human Mesenchymal Stem Cells Reendothelialize Porcine Heart Valve Scaffolds: Novel Perspectives in Heart Valve Tissue Engineering.

Lanuti P, Serafini F, Pierdomenico L, Simeone P, Bologna G, Ercolino E, Di Silvestre S, Guarnieri S, Canosa C, Impicciatore GG, Chiarini S, Magnacca F, Mariggiò MA, Pandolfi A, Marchisio M, Di Giammarco G, Miscia S - Biores Open Access (2015)

Native porcine endothelium observed by fluorescence microscopy and compared with the decellularized porcine scaffold. (A) Both native endothelium (a) and decellularized cusps (c) were stained by DAPI (blue). Autofluorescence of the matrix (green) was observed both on native endothelium (b) and on decellularized cusps (d). (B) Native endothelium was stained by cell membrane labeling PKH26 (red, a) and DAPI (b, blue). A PKH26 and DAPI merge image is also shown (c). Decellularized cusps were stained by cell membrane labeling PKH26 (red, d) and DAPI (e, blue). A PKH26 and DAPI merge image is also shown (f). Images are representative of three separate experiments. Scale bar: 15 μm. DAPI, 4′,6-diamidino-2-phenylindole.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4497625&req=5

f1: Native porcine endothelium observed by fluorescence microscopy and compared with the decellularized porcine scaffold. (A) Both native endothelium (a) and decellularized cusps (c) were stained by DAPI (blue). Autofluorescence of the matrix (green) was observed both on native endothelium (b) and on decellularized cusps (d). (B) Native endothelium was stained by cell membrane labeling PKH26 (red, a) and DAPI (b, blue). A PKH26 and DAPI merge image is also shown (c). Decellularized cusps were stained by cell membrane labeling PKH26 (red, d) and DAPI (e, blue). A PKH26 and DAPI merge image is also shown (f). Images are representative of three separate experiments. Scale bar: 15 μm. DAPI, 4′,6-diamidino-2-phenylindole.
Mentions: To decellularize heart cusps, two different detergent solutions, already reported in the literature, were compared: one composed of 1% SDS+0.05% NaN3,4 and another containing 1% Triton X-100.17,27 To prove the efficacy of the decellularization process, detergent-treated valves were observed by optical microscopy. Figure 1A-c evidences the complete suppression of the porcine endothelium by fluorescence microscopy since nuclei, pointed out by DAPI staining (blue) on the native cusps (Fig. 1A-a), disappeared when decellularized cusps were observed (Fig. 1A-c). In the green channel, the absence of cell nuclei (Fig. 1A-d) underlined the autofluorescence of the decellularized matrix; such a green autofluorescence disappeared when porcine valve are covered by the native endothelium (Fig. 1A-b). Data were confirmed by multiphoton microscopy analyses, showing the absence of the cellular component on the scaffold, after staining the decellularized valves with PKH26 (red) and DAPI (blue) (Fig. 1B-d–f); the native porcine endothelium was used as positive control for nuclei and membrane staining (Fig. 1B-a–c). Of note, the absence of PKH26-positive elements on decellularized cusps revealed that nor cells neither cellular fragments (which may induce immune reactions) were evidenced.

Bottom Line: Here, we were able to successfully decellularize porcine heart valves, which were then recellularized with both differentiated-WJ-MSCs and HUVECs.Cells were able to positively interact with the natural matrix and demonstrated the surface expression of typical endothelial markers.Altogether, these data suggest that the interaction between a biological scaffold and WJ-MSCs allows the regeneration of a morphologically well-structured endothelium, opening new perspectives in the field of HVTE.

View Article: PubMed Central - PubMed

Affiliation: Center for Aging Science (Ce.S.I.), "Università G. d'Annunzio" Foundation , Chieti, Italy . ; Department of Medicine and Aging Science, School of Medicine and Health Science, University "G. d'Annunzio" Chieti-Pescara , Chieti, Italy . ; StemTeCh Group , Chieti, Italy .

ABSTRACT
Heart valve diseases are usually treated by surgical intervention addressed for the replacement of the damaged valve with a biosynthetic or mechanical prosthesis. Although this approach guarantees a good quality of life for patients, it is not free from drawbacks (structural deterioration, nonstructural dysfunction, and reintervention). To overcome these limitations, the heart valve tissue engineering (HVTE) is developing new strategies to synthesize novel types of valve substitutes, by identifying efficient sources of both ideal scaffolds and cells. In particular, a natural matrix, able to interact with cellular components, appears to be a suitable solution. On the other hand, the well-known Wharton's jelly mesenchymal stem cells (WJ-MSCs) plasticity, regenerative abilities, and their immunomodulatory capacities make them highly promising for HVTE applications. In the present study, we investigated the possibility to use porcine valve matrix to regenerate in vitro the valve endothelium by WJ-MSCs differentiated along the endothelial lineage, paralleled with human umbilical vein endothelial cells (HUVECs), used as positive control. Here, we were able to successfully decellularize porcine heart valves, which were then recellularized with both differentiated-WJ-MSCs and HUVECs. Data demonstrated that both cell types were able to reconstitute a cellular monolayer. Cells were able to positively interact with the natural matrix and demonstrated the surface expression of typical endothelial markers. Altogether, these data suggest that the interaction between a biological scaffold and WJ-MSCs allows the regeneration of a morphologically well-structured endothelium, opening new perspectives in the field of HVTE.

No MeSH data available.


Related in: MedlinePlus