Availability of a Magnetic Method for Hepatocyte Transplantation

R. Ito, M. Ito, Y. Asano, A. Murakumo, Naoki Yamamoto, A. Horiguchi

研究成果: Article

抄録

Introduction: Hepatocyte transplantation is a promising alternate for the treatment of hepatic diseases. Hypothermic preservation of isolated human hepatocytes is potentially a simple and convenient strategy to provide on-demand hepatocytes in the quantity sufficient and the quality required for biotherapy. Isolated fresh hepatocytes include damaged cells that are also early apoptotic cells, which is not ideal for hepatocyte transplantation. However, this does not reflect cell viability, although it is considered that it adversely affects cell survival after transplantation. We aimed to harvest these hepatocytes and filter the apoptotic cells using a magnetic method to provide a transplantation source. Materials and Methods: Rat hepatocytes were isolated from caudate lobes using manual enzymatic perfusion. The hepatocyte yield was 5.3 ± 0.66 × 109 cells/g of liver tissue, with a viability of 82.3 ± 3.5%. Two samples of hepatocytes were freshly isolated, one using the magnetic method, and the other without. The magnetic method was performed using DynaMag-15 Magnet, and Annexin V Antibody was used on the early apoptotic cells. We evaluated the viability and plate efficiency of the cells after 24 hours at 37°C. Hepatocytes were isolated using cell separation method, and 30 × 106 cells were mixed with 1.0 mL of Dulbecco's Modified Eagle's Medium (DMEM) and directly injected into the spleen of Lewis rats (150–250 g) using 24-gauge needles. Blood samples were collected on days 0, 3, 7, and 14, and the blood albumin level was measured using enzyme-linked immunosorbent assay (ELISA):G1, control (medium injection); G2, fresh hepatocyte transplant using the magnetic method; and G3, fresh hepatocyte transplant without the magnetic method. Results: The viability was 84.9 ± 2% for fresh hepatocytes and 80.7 ± 1.2% for hepatocytes isolated using the magnetic method. The magnetic method does not damage the cells (73.5 ± 2% vs 35.2 ± 2% after 24 hours), preserving hepatocyte. The albumin level accepted significantly increased in the magnet-treated group compared with the nonmagnet group. Simultaneously, the spleen in which these hepatocytes were transplanted could be used to observe the hepatocytes; the cells were transplanted 14 days later, and the magnet-treated group had significantly higher levels of hepatocytes than the nonmagnet group. Conclusion: We developed an effective technique for hepatocyte isolation for short-term preservation. As a result, we believe that transplantation not only improves the cell transplantation effect but also allows the cells to be stored efficiently using the magnetic method. These results demonstrate the usefulness of hepatocyte hypothermic preservation for cell transplantation.

元の言語English
ページ(範囲)1525-1531
ページ数7
ジャーナルTransplantation Proceedings
50
発行部数5
DOI
出版物ステータスPublished - 01-06-2018

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Hepatocytes
Transplantation
Magnets
Cell Transplantation
Albumins
Cell Survival
Spleen
Transplants
Eagles
Biological Therapy
Cell Separation
Annexin A5
Liver
Needles

All Science Journal Classification (ASJC) codes

  • Surgery
  • Transplantation

これを引用

Ito, R. ; Ito, M. ; Asano, Y. ; Murakumo, A. ; Yamamoto, Naoki ; Horiguchi, A. / Availability of a Magnetic Method for Hepatocyte Transplantation. :: Transplantation Proceedings. 2018 ; 巻 50, 番号 5. pp. 1525-1531.
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abstract = "Introduction: Hepatocyte transplantation is a promising alternate for the treatment of hepatic diseases. Hypothermic preservation of isolated human hepatocytes is potentially a simple and convenient strategy to provide on-demand hepatocytes in the quantity sufficient and the quality required for biotherapy. Isolated fresh hepatocytes include damaged cells that are also early apoptotic cells, which is not ideal for hepatocyte transplantation. However, this does not reflect cell viability, although it is considered that it adversely affects cell survival after transplantation. We aimed to harvest these hepatocytes and filter the apoptotic cells using a magnetic method to provide a transplantation source. Materials and Methods: Rat hepatocytes were isolated from caudate lobes using manual enzymatic perfusion. The hepatocyte yield was 5.3 ± 0.66 × 109 cells/g of liver tissue, with a viability of 82.3 ± 3.5{\%}. Two samples of hepatocytes were freshly isolated, one using the magnetic method, and the other without. The magnetic method was performed using DynaMag-15 Magnet, and Annexin V Antibody was used on the early apoptotic cells. We evaluated the viability and plate efficiency of the cells after 24 hours at 37°C. Hepatocytes were isolated using cell separation method, and 30 × 106 cells were mixed with 1.0 mL of Dulbecco's Modified Eagle's Medium (DMEM) and directly injected into the spleen of Lewis rats (150–250 g) using 24-gauge needles. Blood samples were collected on days 0, 3, 7, and 14, and the blood albumin level was measured using enzyme-linked immunosorbent assay (ELISA):G1, control (medium injection); G2, fresh hepatocyte transplant using the magnetic method; and G3, fresh hepatocyte transplant without the magnetic method. Results: The viability was 84.9 ± 2{\%} for fresh hepatocytes and 80.7 ± 1.2{\%} for hepatocytes isolated using the magnetic method. The magnetic method does not damage the cells (73.5 ± 2{\%} vs 35.2 ± 2{\%} after 24 hours), preserving hepatocyte. The albumin level accepted significantly increased in the magnet-treated group compared with the nonmagnet group. Simultaneously, the spleen in which these hepatocytes were transplanted could be used to observe the hepatocytes; the cells were transplanted 14 days later, and the magnet-treated group had significantly higher levels of hepatocytes than the nonmagnet group. Conclusion: We developed an effective technique for hepatocyte isolation for short-term preservation. As a result, we believe that transplantation not only improves the cell transplantation effect but also allows the cells to be stored efficiently using the magnetic method. These results demonstrate the usefulness of hepatocyte hypothermic preservation for cell transplantation.",
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Availability of a Magnetic Method for Hepatocyte Transplantation. / Ito, R.; Ito, M.; Asano, Y.; Murakumo, A.; Yamamoto, Naoki; Horiguchi, A.

:: Transplantation Proceedings, 巻 50, 番号 5, 01.06.2018, p. 1525-1531.

研究成果: Article

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T1 - Availability of a Magnetic Method for Hepatocyte Transplantation

AU - Ito, R.

AU - Ito, M.

AU - Asano, Y.

AU - Murakumo, A.

AU - Yamamoto, Naoki

AU - Horiguchi, A.

PY - 2018/6/1

Y1 - 2018/6/1

N2 - Introduction: Hepatocyte transplantation is a promising alternate for the treatment of hepatic diseases. Hypothermic preservation of isolated human hepatocytes is potentially a simple and convenient strategy to provide on-demand hepatocytes in the quantity sufficient and the quality required for biotherapy. Isolated fresh hepatocytes include damaged cells that are also early apoptotic cells, which is not ideal for hepatocyte transplantation. However, this does not reflect cell viability, although it is considered that it adversely affects cell survival after transplantation. We aimed to harvest these hepatocytes and filter the apoptotic cells using a magnetic method to provide a transplantation source. Materials and Methods: Rat hepatocytes were isolated from caudate lobes using manual enzymatic perfusion. The hepatocyte yield was 5.3 ± 0.66 × 109 cells/g of liver tissue, with a viability of 82.3 ± 3.5%. Two samples of hepatocytes were freshly isolated, one using the magnetic method, and the other without. The magnetic method was performed using DynaMag-15 Magnet, and Annexin V Antibody was used on the early apoptotic cells. We evaluated the viability and plate efficiency of the cells after 24 hours at 37°C. Hepatocytes were isolated using cell separation method, and 30 × 106 cells were mixed with 1.0 mL of Dulbecco's Modified Eagle's Medium (DMEM) and directly injected into the spleen of Lewis rats (150–250 g) using 24-gauge needles. Blood samples were collected on days 0, 3, 7, and 14, and the blood albumin level was measured using enzyme-linked immunosorbent assay (ELISA):G1, control (medium injection); G2, fresh hepatocyte transplant using the magnetic method; and G3, fresh hepatocyte transplant without the magnetic method. Results: The viability was 84.9 ± 2% for fresh hepatocytes and 80.7 ± 1.2% for hepatocytes isolated using the magnetic method. The magnetic method does not damage the cells (73.5 ± 2% vs 35.2 ± 2% after 24 hours), preserving hepatocyte. The albumin level accepted significantly increased in the magnet-treated group compared with the nonmagnet group. Simultaneously, the spleen in which these hepatocytes were transplanted could be used to observe the hepatocytes; the cells were transplanted 14 days later, and the magnet-treated group had significantly higher levels of hepatocytes than the nonmagnet group. Conclusion: We developed an effective technique for hepatocyte isolation for short-term preservation. As a result, we believe that transplantation not only improves the cell transplantation effect but also allows the cells to be stored efficiently using the magnetic method. These results demonstrate the usefulness of hepatocyte hypothermic preservation for cell transplantation.

AB - Introduction: Hepatocyte transplantation is a promising alternate for the treatment of hepatic diseases. Hypothermic preservation of isolated human hepatocytes is potentially a simple and convenient strategy to provide on-demand hepatocytes in the quantity sufficient and the quality required for biotherapy. Isolated fresh hepatocytes include damaged cells that are also early apoptotic cells, which is not ideal for hepatocyte transplantation. However, this does not reflect cell viability, although it is considered that it adversely affects cell survival after transplantation. We aimed to harvest these hepatocytes and filter the apoptotic cells using a magnetic method to provide a transplantation source. Materials and Methods: Rat hepatocytes were isolated from caudate lobes using manual enzymatic perfusion. The hepatocyte yield was 5.3 ± 0.66 × 109 cells/g of liver tissue, with a viability of 82.3 ± 3.5%. Two samples of hepatocytes were freshly isolated, one using the magnetic method, and the other without. The magnetic method was performed using DynaMag-15 Magnet, and Annexin V Antibody was used on the early apoptotic cells. We evaluated the viability and plate efficiency of the cells after 24 hours at 37°C. Hepatocytes were isolated using cell separation method, and 30 × 106 cells were mixed with 1.0 mL of Dulbecco's Modified Eagle's Medium (DMEM) and directly injected into the spleen of Lewis rats (150–250 g) using 24-gauge needles. Blood samples were collected on days 0, 3, 7, and 14, and the blood albumin level was measured using enzyme-linked immunosorbent assay (ELISA):G1, control (medium injection); G2, fresh hepatocyte transplant using the magnetic method; and G3, fresh hepatocyte transplant without the magnetic method. Results: The viability was 84.9 ± 2% for fresh hepatocytes and 80.7 ± 1.2% for hepatocytes isolated using the magnetic method. The magnetic method does not damage the cells (73.5 ± 2% vs 35.2 ± 2% after 24 hours), preserving hepatocyte. The albumin level accepted significantly increased in the magnet-treated group compared with the nonmagnet group. Simultaneously, the spleen in which these hepatocytes were transplanted could be used to observe the hepatocytes; the cells were transplanted 14 days later, and the magnet-treated group had significantly higher levels of hepatocytes than the nonmagnet group. Conclusion: We developed an effective technique for hepatocyte isolation for short-term preservation. As a result, we believe that transplantation not only improves the cell transplantation effect but also allows the cells to be stored efficiently using the magnetic method. These results demonstrate the usefulness of hepatocyte hypothermic preservation for cell transplantation.

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