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Human saphenous vein organ culture under controlled hemodynamic conditions.

Miyakawa AA, Dallan LA, Lacchini S, Borin TF, Krieger JE - Clinics (Sao Paulo) (2008)

Bottom Line: Cell viability, cell density and apoptosis were compared before and after these intervals using MTT, Hoeschst 33258 stain, and TUNEL assays, respectively.Although the cell density from 1-day cultured veins under arterial conditions was similar to that of freshly excised veins, the TUNEL assay indicated that most of these cells were undergoing apoptosis.The results observed resemble the events taking place during early in vivo arterial-vein grafting and provide evidence that an ex vivo perfusion system may be useful for the identification of new therapeutic targets that ameliorate vein graft remodeling and increase graft patency over time.

View Article: PubMed Central - PubMed

Affiliation: Heart Institute (InCor) and Department of Medicine-LIM 13, Faculdade de Medicina, Universidade de São Paulo, São Paulo, SP, Brazil. ayumi.miyakawa@incor.usp.br

ABSTRACT

Introduction: Saphenous vein grafting is still widely used to revascularize ischemic myocardium. The effectiveness of this procedure is limited by neointima formation and accelerated atherosclerosis, which frequently leads to graft occlusion. A better understanding of this process is important to clarify the mechanisms of vein graft disease and to aid in the formulation of strategies for prevention and/or therapeutics.

Objective: To develop an ex vivo flow system that allows for controlled hemodynamics in order to mimic arterial and venous conditions.

Methods: Human saphenous veins were cultured either under venous (flow: 5 ml/min) or arterial hemodynamic conditions (flow: 50 ml/min, pressure: 80 mmHg) for 1-, 2- and 4-day periods. Cell viability, cell density and apoptosis were compared before and after these intervals using MTT, Hoeschst 33258 stain, and TUNEL assays, respectively.

Results: Fresh excised tissue segments were well preserved prior to the study. Hoechst 33258 and MTT stains showed progressive losses in cell density and cell viability in veins cultured under arterial hemodynamic conditions from 1 to 4 days, while no alterations were observed in veins cultured under venous conditions. Although the cell density from 1-day cultured veins under arterial conditions was similar to that of freshly excised veins, the TUNEL assay indicated that most of these cells were undergoing apoptosis.

Conclusion: The results observed resemble the events taking place during early in vivo arterial-vein grafting and provide evidence that an ex vivo perfusion system may be useful for the identification of new therapeutic targets that ameliorate vein graft remodeling and increase graft patency over time.

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Related in: MedlinePlus

Hoechst 33258 nuclei stain of human saphenous vein cultured in the ex vivo perfusion system. The segments were cultured under venous (A, C, E) and arterial (B, D, F) hemodynamic conditions for 1 (A, B), 2 (C, D) or 4 (E, F) days. 100X magnification
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f3-18-0106: Hoechst 33258 nuclei stain of human saphenous vein cultured in the ex vivo perfusion system. The segments were cultured under venous (A, C, E) and arterial (B, D, F) hemodynamic conditions for 1 (A, B), 2 (C, D) or 4 (E, F) days. 100X magnification

Mentions: The HE staining was also performed in vein segments cultured under venous and arterial conditions, and the number of nuclei stained by hematoxylin gradually diminished from 1 to 4 days when cultured in the arterial condition. No alteration was observed in veins cultured under venous conditions (data not shown). To further confirm the HE staining results, Hoechst 33258 nuclei staining was used to verify cell density in the vessel segments cultured under venous and arterial stimuli. As noted before, cellular density tended to decrease in specimens cultured under arterial conditions for 4 days, whereas no modification was observed in the veins cultured under venous conditions (Figures 3 and 4).


Human saphenous vein organ culture under controlled hemodynamic conditions.

Miyakawa AA, Dallan LA, Lacchini S, Borin TF, Krieger JE - Clinics (Sao Paulo) (2008)

Hoechst 33258 nuclei stain of human saphenous vein cultured in the ex vivo perfusion system. The segments were cultured under venous (A, C, E) and arterial (B, D, F) hemodynamic conditions for 1 (A, B), 2 (C, D) or 4 (E, F) days. 100X magnification
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2664728&req=5

f3-18-0106: Hoechst 33258 nuclei stain of human saphenous vein cultured in the ex vivo perfusion system. The segments were cultured under venous (A, C, E) and arterial (B, D, F) hemodynamic conditions for 1 (A, B), 2 (C, D) or 4 (E, F) days. 100X magnification
Mentions: The HE staining was also performed in vein segments cultured under venous and arterial conditions, and the number of nuclei stained by hematoxylin gradually diminished from 1 to 4 days when cultured in the arterial condition. No alteration was observed in veins cultured under venous conditions (data not shown). To further confirm the HE staining results, Hoechst 33258 nuclei staining was used to verify cell density in the vessel segments cultured under venous and arterial stimuli. As noted before, cellular density tended to decrease in specimens cultured under arterial conditions for 4 days, whereas no modification was observed in the veins cultured under venous conditions (Figures 3 and 4).

Bottom Line: Cell viability, cell density and apoptosis were compared before and after these intervals using MTT, Hoeschst 33258 stain, and TUNEL assays, respectively.Although the cell density from 1-day cultured veins under arterial conditions was similar to that of freshly excised veins, the TUNEL assay indicated that most of these cells were undergoing apoptosis.The results observed resemble the events taking place during early in vivo arterial-vein grafting and provide evidence that an ex vivo perfusion system may be useful for the identification of new therapeutic targets that ameliorate vein graft remodeling and increase graft patency over time.

View Article: PubMed Central - PubMed

Affiliation: Heart Institute (InCor) and Department of Medicine-LIM 13, Faculdade de Medicina, Universidade de São Paulo, São Paulo, SP, Brazil. ayumi.miyakawa@incor.usp.br

ABSTRACT

Introduction: Saphenous vein grafting is still widely used to revascularize ischemic myocardium. The effectiveness of this procedure is limited by neointima formation and accelerated atherosclerosis, which frequently leads to graft occlusion. A better understanding of this process is important to clarify the mechanisms of vein graft disease and to aid in the formulation of strategies for prevention and/or therapeutics.

Objective: To develop an ex vivo flow system that allows for controlled hemodynamics in order to mimic arterial and venous conditions.

Methods: Human saphenous veins were cultured either under venous (flow: 5 ml/min) or arterial hemodynamic conditions (flow: 50 ml/min, pressure: 80 mmHg) for 1-, 2- and 4-day periods. Cell viability, cell density and apoptosis were compared before and after these intervals using MTT, Hoeschst 33258 stain, and TUNEL assays, respectively.

Results: Fresh excised tissue segments were well preserved prior to the study. Hoechst 33258 and MTT stains showed progressive losses in cell density and cell viability in veins cultured under arterial hemodynamic conditions from 1 to 4 days, while no alterations were observed in veins cultured under venous conditions. Although the cell density from 1-day cultured veins under arterial conditions was similar to that of freshly excised veins, the TUNEL assay indicated that most of these cells were undergoing apoptosis.

Conclusion: The results observed resemble the events taking place during early in vivo arterial-vein grafting and provide evidence that an ex vivo perfusion system may be useful for the identification of new therapeutic targets that ameliorate vein graft remodeling and increase graft patency over time.

Show MeSH
Related in: MedlinePlus