Transplantation of Human induced pluripotent stem cell (iPSC)-derived hepatocyte-like cells (iPS-H) under the murine kidney and liver capsule
1.The purpose of the trial
Liver transplantation is a successful treatment option for patients with end-stage liver disease. However, transplantable donor livers are in short supply and maturation of human induced pluripotent stem cells (hiPSCs) to hepatocyte-like cells (HLCs) is proposed to address the limitation of human hepatocytes for therapeutic applications.
Human induced pluripotent stem cell (iPSC)-derived hepatocyte-like cells (iPS-H) hold tremendous promise for regenerative medicine, toxicology screening, and developmental biology, but a major challenge is to produce hepatocyte like cells (HLCs) from pluripotent stem cells (PSCs) that faithfully recapitulate primary human hepatocytes in terms of their metabolic potential.
The Gareth Sullivan Lab have developed a procedure to differentiate hiPSC lines to functional hepatocyte organoids which exhibit basal metabolism, production of serum proteins and key markers such as albumin in vitro (Sullivan et al., 2009, Siller et al., 2017). In collaboration with the Gareth Sullivan Lab, we now aim to perform a proof of principle study by investigating whether these iPS-H also are able to produce human albumin in vivo.
2. Adverse effects:
Mild/slightly stressful (mild belastningsgrad).
We expect the animals to experience a transient mild pain post surgery.
We do not expect any impairment of the animals welfare or general condition.
3. Expected utility
If we are able to prove that the iPS-H-cells also are able to produce human albumin in vivo (and not just in vitro in a dish) it means that these cells have passed the second quality control check where they are able to function as human hepatocytes within a living organism, and we are one step closer to address the limitation of human hepatocytes for therapeutic application.
4. Number of animals and animal species: Mus musculus, 36 animals in total (27 mice subjected to experiment, 9 mice in back up in case of error).
In order to do this we will subject 8-10 weeks old male immunodeficient 001303 - NOD.CB17-Prkdcscid/J mice from Jackson (3 in each group) to three different methods of iPS-H injection; iPS-H suspension transplantation under kidney capsule, iPS-H suspension transplantation under liver capsule or intraperitioneal injection (I.P).The I.P treatment is included to ensure success,while the kidney is easily injectable. From the 1st week and the following 2 months, we will weekly draw 10ul blood from the saphena vein and measure human albumin by ELISA.
Import and in vivo use of biological material in accordance with OUS procedure 114241, attachment 1. The hiPSC line cell for engraftment was reprogrammed from AG05836 from the Coriell Institute (https://www.coriell.org/0/Sections/Search/Sample_Detail.aspx?Ref=AG05836&PgId=166) in the Gareth Sullivan Lab.
5. How to comply with 3R:
The replacement option is already performed.
The n-value is 3 in each group, the lowest possible number to ensure trustworthy repeatability
Since this is a proof of principle study, no control animals is necessary.The surgery is based upon the internal experience and experience from a large number of research groups, as this method is highly well established. Pain relief is included.
Liver transplantation is a successful treatment option for patients with end-stage liver disease. However, transplantable donor livers are in short supply and maturation of human induced pluripotent stem cells (hiPSCs) to hepatocyte-like cells (HLCs) is proposed to address the limitation of human hepatocytes for therapeutic applications.
Human induced pluripotent stem cell (iPSC)-derived hepatocyte-like cells (iPS-H) hold tremendous promise for regenerative medicine, toxicology screening, and developmental biology, but a major challenge is to produce hepatocyte like cells (HLCs) from pluripotent stem cells (PSCs) that faithfully recapitulate primary human hepatocytes in terms of their metabolic potential.
The Gareth Sullivan Lab have developed a procedure to differentiate hiPSC lines to functional hepatocyte organoids which exhibit basal metabolism, production of serum proteins and key markers such as albumin in vitro (Sullivan et al., 2009, Siller et al., 2017). In collaboration with the Gareth Sullivan Lab, we now aim to perform a proof of principle study by investigating whether these iPS-H also are able to produce human albumin in vivo.
2. Adverse effects:
Mild/slightly stressful (mild belastningsgrad).
We expect the animals to experience a transient mild pain post surgery.
We do not expect any impairment of the animals welfare or general condition.
3. Expected utility
If we are able to prove that the iPS-H-cells also are able to produce human albumin in vivo (and not just in vitro in a dish) it means that these cells have passed the second quality control check where they are able to function as human hepatocytes within a living organism, and we are one step closer to address the limitation of human hepatocytes for therapeutic application.
4. Number of animals and animal species: Mus musculus, 36 animals in total (27 mice subjected to experiment, 9 mice in back up in case of error).
In order to do this we will subject 8-10 weeks old male immunodeficient 001303 - NOD.CB17-Prkdcscid/J mice from Jackson (3 in each group) to three different methods of iPS-H injection; iPS-H suspension transplantation under kidney capsule, iPS-H suspension transplantation under liver capsule or intraperitioneal injection (I.P).The I.P treatment is included to ensure success,while the kidney is easily injectable. From the 1st week and the following 2 months, we will weekly draw 10ul blood from the saphena vein and measure human albumin by ELISA.
Import and in vivo use of biological material in accordance with OUS procedure 114241, attachment 1. The hiPSC line cell for engraftment was reprogrammed from AG05836 from the Coriell Institute (https://www.coriell.org/0/Sections/Search/Sample_Detail.aspx?Ref=AG05836&PgId=166) in the Gareth Sullivan Lab.
5. How to comply with 3R:
The replacement option is already performed.
The n-value is 3 in each group, the lowest possible number to ensure trustworthy repeatability
Since this is a proof of principle study, no control animals is necessary.The surgery is based upon the internal experience and experience from a large number of research groups, as this method is highly well established. Pain relief is included.