Efficient Differentiation of Human Pluripotent Stem Cells into Liver Cells

Kyle  M. Loh; Amrita Palaria; Lay Teng Ang

doi:10.3791/58975

JoVE Journal
Developmental Biology

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JoVE Journal Developmental Biology

Efficient Differentiation of Human Pluripotent Stem Cells into Liver Cells

인간 다능성 줄기 세포를 간 세포로 효율적으로 분화

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07:37 min

June 11, 2019

DOI: 10.3791/58975-v

Kyle M. Loh^1,2, Amrita Palaria¹, Lay Teng Ang¹

¹Institute for Stem Cell Biology & Regenerative Medicine, Stanford-UC Berkeley Siebel Stem Cell Institute,Stanford University School of Medicine, ²Department of Developmental Biology,Stanford University School of Medicine

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Overview

This protocol describes a serum-free method for generating hepatocyte-like cells from human pluripotent stem cells (hPSCs) in just 18 days. The process involves six steps of differentiation through various intermediate cell types.

Key Study Components

Area of Science

Stem Cell Biology
Regenerative Medicine
Cell Differentiation

Background

Human liver cells are crucial for studying liver diseases and drug metabolism.
Current methods for generating liver cells often yield low purity.
Efficient generation of hepatocyte-like cells can advance regenerative therapies.
This protocol aims to improve the purity and yield of liver progenitor cells.

Purpose of Study

To develop a high-purity method for generating hepatocyte-like cells from hPSCs.
To enhance the understanding of liver cell differentiation pathways.
To provide a reliable source of liver cells for research and therapeutic applications.

Methods Used

Sequential differentiation of hPSCs through defined stages.
Control of developmental signaling pathways to promote liver differentiation.
Use of serum-free culture conditions to enhance cell purity.
Preparation of PVA stock for medium formulation.

Main Results

High purity of hepatocyte-like cells achieved within 18 days.
Efficient generation of liver bud progenitors.
Demonstrated control over unwanted cell type formation.
Potential applications in regenerative medicine highlighted.

Conclusions

This method provides a reliable approach to generate liver cells from hPSCs.
High purity and yield can facilitate research in liver biology.
Future studies may explore therapeutic applications of these cells.

Frequently Asked Questions

What are hepatocyte-like cells?

Hepatocyte-like cells are cells that mimic the function and characteristics of liver cells, derived from stem cells.

How long does the differentiation process take?

The differentiation process takes 18 days to generate hepatocyte-like cells.

What is the significance of using serum-free conditions?

Serum-free conditions help improve the purity of the generated liver cells and reduce variability.

Can this method be applied to other cell types?

While this method is specific for liver cells, similar techniques may be adapted for other cell types.

What are the potential applications of these hepatocyte-like cells?

They can be used in drug testing, disease modeling, and regenerative medicine.

What are the main challenges in generating liver cells from hPSCs?

Challenges include achieving high purity and controlling differentiation pathways effectively.

이 프로토콜은 18일 만에 인간 만능 줄기 세포(hPSCs)로부터 간세포 유사 세포를 효율적으로 생성하는 단층, 무혈청 방법을 상세히 기술한다. 이것은 hPSCs가 간세포 같이 세포를 형성하기 전에 원시적인 선미, 결정적인 내배엽, 후방 포인및 간 싹 선조와 같은 중간 세포 모형으로 순차적으로 분화하는 것과 같이 6단계를 수반합니다.

이 방법은 높은 순도인간 간싹 선조및 간세포 같이 세포의 다수의 생성을 가능하게 합니다. 이 간 세포를 생성하는 기능은 재생 의학 및 그밖 필드에 중요할 수 있었습니다. 간 분화를 촉진하고 원치 않는 세포 유형의 형성을 억제하기 위해 발달 신호 경로를 제어함으로써,이 방법은 각각 분화의 6 및 18 일에 의해 인간 간 새싹 선조및 간세포 와 같은 세포의 효율적인 생성을 가능하게한다.

이 기술의 주요 장점은 생성 된 인간 간 세포의 높은 순도이다. 이 절차이기 위해 교반 바가 들어 있는 원추형 플라스크에 50밀리리터의 IMDM을 추가합니다. 중간 을 섭씨 50도까지 가열하고 0.5 그램의 PVA를 추가하면서 지속적으로 교반하여 밀리리터 당 10 밀리그램의 농도로 PVA 스톡을 준비합니다.

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