Preliminary experience with tissue engineering of a venous vascular patch by using bone marrow–derived cells and a hybrid biodegradable polymer scaffold

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Presentation transcript:

Preliminary experience with tissue engineering of a venous vascular patch by using bone marrow–derived cells and a hybrid biodegradable polymer scaffold Seung-Woo Cho, PhD, Oju Jeon, MS, Joung Eun Lim, MS, So-Jung Gwak, MS, Sang-Soo Kim, PhD, Cha Yong Choi, PhD, Dong-Ik Kim, MD, Byung-Soo Kim, PhD Journal of Vascular Surgery Volume 44, Issue 6, Pages 1329-1340 (December 2006) DOI: 10.1016/j.jvs.2006.07.032 Copyright © 2006 The Society for Vascular Surgery Terms and Conditions

Fig 1 Vascular patch scaffold fabricated from poly(lactide-co-ε-caprolactone) (PLCL) copolymer reinforced with poly(glycolic acid) (PGA) fibers. (A) Gross view of a PGA/PLCL patch scaffold (15 mm wide × 30 mm long). The scale is in centimeters. Scanning electron micrographs of (B) cross section (original magnification, ×100), (C) luminal surface (original magnification, ×200), and (D) external surface (original magnification, ×70) of PGA/PLCL patch scaffold are shown. Scale bars = 200 μm. Journal of Vascular Surgery 2006 44, 1329-1340DOI: (10.1016/j.jvs.2006.07.032) Copyright © 2006 The Society for Vascular Surgery Terms and Conditions

Fig 2 Tensile mechanical properties of native blood vessels (canine inferior venae cavae [IVCs]; n = 3) and poly(glycolic acid) (PGA)/poly(lactide-co-ε-caprolactone) (PLCL) (n = 3) and PLCL scaffolds (n = 3) after incubation in phosphate-buffered saline with shaking at 37°C for various time periods. A, Tensile strength of PGA/PLCL scaffolds was significantly higher (P < .05) than that of PLCL scaffolds at 0, 1, 2, and 8 weeks after incubation. B, Tensile elongation of PGA/PLCL scaffolds was significantly lower (P < .05) than that of PLCL scaffolds at 0, 1, 2, and 4 weeks after incubation. At 4 and 8 weeks after incubation, tensile elongation of PGA/PLCL scaffolds was not significantly different (P > .05) from that of native blood vessels. Journal of Vascular Surgery 2006 44, 1329-1340DOI: (10.1016/j.jvs.2006.07.032) Copyright © 2006 The Society for Vascular Surgery Terms and Conditions

Fig 3 Characterization of differentiated bone marrow–derived cells. (A) Endothelial cells (ECs) derived from bone marrow mononuclear cells (BMMNCs) showed a cobblestone morphology (original magnification, ×100) and stained positively for (B) von Willebrand factor (vWF; original magnification, ×400) and (C) CD31 (original magnification, ×400). In control staining without the use of primary antibody, ECs derived from BMMNCs stained negatively for (D) vWF (original magnification, ×400). (E) Smooth muscle cells (SMCs) derived from BMMNCs showed the morphology of mature SMCs (original magnification, ×200) and stained positively for (F) smooth muscle (SM) α-actin (original magnification, ×200) and (G) SM myosin heavy chain (original magnification, ×200). In control staining without the use of primary antibody, SMCs derived from BMMNCs stained negatively for (H) SM α-actin (original magnification, ×200). Scale bars = 30 μm. Journal of Vascular Surgery 2006 44, 1329-1340DOI: (10.1016/j.jvs.2006.07.032) Copyright © 2006 The Society for Vascular Surgery Terms and Conditions

Fig 4 Preimplantation examinations of bone marrow–derived cell (BMC)-seeded vascular patches. (A) Hematoxylin and eosin–stained section (original magnification, ×400) of tissue-engineered vascular patch maintained in vitro for 1 week after cell seeding [black arrow, poly(glycolic acid) fiber; black arrowheads, poly(lactide-co-ε-caprolactone); white arrows, cellular portion]. Scale bar = 100 μm. (B) Scanning electron micrograph of the luminal surface of a vascular patch 1 week after BMC seeding (original magnification, ×600). Scale bar = 30 μm. Immunohistochemical staining is shown of cell-seeded vascular patches for (C) von Willebrand factor (original magnification, ×400; scale bar = 100 μm) and (D) smooth muscle α-actin (original magnification, ×400; scale bar = 100 μm). (E) Chloromethyl-1,1′-dioactadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate–labeled cells in the patch scaffolds 1 week after cell seeding (original magnification, ×200) are shown. Scale bar = 200 μm. Journal of Vascular Surgery 2006 44, 1329-1340DOI: (10.1016/j.jvs.2006.07.032) Copyright © 2006 The Society for Vascular Surgery Terms and Conditions

Fig 5 Implantation and explantation of tissue-engineered vascular patches. A, Gross view of a poly(glycolic acid)/poly(lactide-co-ε-caprolactone) vascular patch seeded with bone marrow–derived cells (BMCs) before implantation. The scale is in centimeters. B, Surgical implantation of tissue-engineered vascular patches. The vascular patch replaced the inferior vena cava (IVC) segments of bone marrow donor dogs. C, Gross view of a vascular patch retrieved 8 weeks after implantation. A vascular patch engineered with BMCs showed a smooth luminal surface without any sign of thrombosis. Arrows indicate the vascular patch implanted into the IVC. The scale is in centimeters. Journal of Vascular Surgery 2006 44, 1329-1340DOI: (10.1016/j.jvs.2006.07.032) Copyright © 2006 The Society for Vascular Surgery Terms and Conditions

Fig 6 Histologic and immunohistochemical analyses of vascular patches retrieved 8 weeks after implantation. A, Hematoxylin and eosin staining of retrieved patches showed regeneration of vascular tissues (original magnification, ×100; scale bar = 200 μm). B, Masson trichrome staining indicated collagen regeneration (original magnification, ×100; scale bar = 200 μm). C, Cells on the luminal sides of retrieved patches stained positively for von Willebrand factor, thus indicating endothelium regeneration (original magnification, ×400; scale bar = 100 μm). D, Immunohistochemical staining for smooth muscle (SM) α-actin (original magnification, ×400; scale bar = 100 μm) showed regeneration of SM tissue. Journal of Vascular Surgery 2006 44, 1329-1340DOI: (10.1016/j.jvs.2006.07.032) Copyright © 2006 The Society for Vascular Surgery Terms and Conditions

Fig 7 Scanning electron microscopic examinations for confirmation of endothelium. A, The endothelium regenerated in the tissue-engineered vascular patches retrieved 8 weeks after implantation (original magnification, ×1500). B, The endothelium of native canine inferior vena cava (IVC; original magnification, ×1500). These images are the representative images of the luminal surface of tissue-engineered vascular patches and native IVCs. Scale bars = 20 μm. Journal of Vascular Surgery 2006 44, 1329-1340DOI: (10.1016/j.jvs.2006.07.032) Copyright © 2006 The Society for Vascular Surgery Terms and Conditions

Fig 8 Examination of calcification in vascular patches retrieved 8 weeks after implantation. (A) Calcium contents of tissue-engineered vascular patches at 8 weeks after implantation and of native canine inferior venae cavae (IVCs). The calcium content of tissue-engineered vascular patches (n = 4) was not statistically significantly different (P > .05) from that of native IVCs (n = 4). Immunohistochemical staining of (B) tissue-engineered vascular patches (original magnification, ×100), (C) native IVCs (original magnification, ×100), and (D) native canine bone tissue (original magnification, ×400) for osteopontin is shown. Scale bars = 100 μm. Journal of Vascular Surgery 2006 44, 1329-1340DOI: (10.1016/j.jvs.2006.07.032) Copyright © 2006 The Society for Vascular Surgery Terms and Conditions

Fig 9 Immunofluorescent double staining of tissue-engineered vascular patches retrieved 8 weeks after implantation. A, Immunofluorescent staining for von Willebrand factor (vWF) showed endothelium (green) formation on the luminal sides of the patches (original magnification, ×200; scale bar = 100 μm). B, Cells labeled with chloromethyl-1,1′-dioactadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate (CM-DiI; red) before implantation were present in the luminal sides of patches retrieved at 8 weeks (original magnification, ×200; scale bar = 100 μm). C, Merged image of vWF-positive cells and CM-DiI–labeled cells (original magnification, ×200; scale bar = 100 μm). D, Immunofluorescent staining for smooth muscle (SM) α-actin showed SM (green) regeneration in the medial regions of patches (original magnification, ×400; scale bar = 50 μm). E, CM-DiI–labeled cells (red) were present in the medial parts of retrieved patches (original magnification, ×400; scale bar = 50 μm). F, Merged image of SM α-actin–positive cells and CM-DiI–labeled cells (original magnification, ×400; scale bar = 50 μm). Journal of Vascular Surgery 2006 44, 1329-1340DOI: (10.1016/j.jvs.2006.07.032) Copyright © 2006 The Society for Vascular Surgery Terms and Conditions